Network-wide abnormalities explain memory variability in hippocampal amnesia

Abstract
Introduction
Results
Discussion
Materials and methods
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Abstract

Patients with hippocampal amnesia play a central role in memory neuroscience but the neural underpinnings of amnesia are hotly debated. We hypothesized that focal hippocampal damage is associated with changes across the extended hippocampal system and that these, rather than hippocampal atrophy per se, would explain variability in memory between patients. We assessed this hypothesis in a uniquely large cohort of patients (n = 38) after autoimmune limbic encephalitis, a syndrome associated with focal structural hippocampal pathology. These patients showed impaired recall, recognition and maintenance of new information, and remote autobiographical amnesia. Besides hippocampal atrophy, we observed correlatively reduced thalamic and entorhinal cortical volume, resting-state inter-hippocampal connectivity and activity in posteromedial cortex. Associations of hippocampal volume with recall, recognition, and remote memory were fully mediated by wider network abnormalities, and were only direct in forgetting. Network abnormalities may explain the variability across studies of amnesia and speak to debates in memory neuroscience.

https://doi.org/10.7554/eLife.46156.001

Introduction

Ever since the first report of the famous patient H.M. (Scoville and Milner, 1957), hippocampal (HPC) amnesia has played a fundamental role in the neuroscience of human memory (MacPherson and Della Sala, 2019). Studies of amnesic patients have, however, often produced inconsistent results. Such studies have been criticized for attributing behavioral deficits solely to the HPC (Squire and Wixted, 2011), despite the fact that focal damage may trigger more subtle structural and functional abnormalities across an extended HPC system, involving the thalamus and cingulate cortex (Aggleton, 2014). Indeed, broader network abnormalities are well documented in other ‘focal’ conditions [e.g. ischemic stroke (Carter et al., 2010; Veldsman et al., 2018)] and have been demonstrated in small case series of amnesic patients (Hayes et al., 2012; Henson et al., 2016; Rudebeck et al., 2013). These abnormalities may be important in understanding memory impairment in neurological disease (Addis et al., 2007) but their explanatory potential is under-explored.

We propose that network abnormalities are key to understanding inconsistencies that underpin central debates about HPC function in amnesia research. One such debate centers on the dissociation of neurocognitive processes underlying fundamental aspects of anterograde retrieval. Some authors hold that recall relies on HPC-dependent recollection processes, whereas recognition additionally draws on familiarity, which is mediated by non-HPC structures within the medial temporal lobe (MTL) (Brown and Aggleton, 2001; Kafkas et al., 2016). Others argue that recollection and familiarity do not dissociate within the MTL (Wixted and Squire, 2010). Related to the above, some theorists posit that HPC processing is material-specific, whereby the HPC holds a privileged role in processing scenes or spatial stimuli and this function underpins its involvement in episodic memory and navigation (Maguire and Mullally, 2013; Graham et al., 2010). An opposing view holds that the primary role of the HPC is the material-independent maintenance of information over a limited time period (Kim et al., 2015). A second, long-standing debate (Cassel and Kopelman, 2019), pertains to the presence of abnormal (accelerated) forgetting in HPC amnesia. Whereas some reports have provided evidence for abnormal forgetting after MTL damage (Huppert and Piercy, 1979; Isaac and Mayes, 1999), other studies have reported normal forgetting rates (Freed et al., 1987; Kopelman, 1985; McKee and Squire, 1992). A third debate pertains to the HPC role in remote memory. The ‘standard consolidation theory’ holds that newly-acquired memories initially depend upon the HPC but that, through a process of ‘consolidation’, memory traces gradually become HPC-independent and are stored in and retrieved by neocortical structures alone (Hardt and Nadel, 2018; Squire et al., 2015). In contrast, ‘multiple trace theory’ proposes that the HPC is necessary for vivid episodic recollection irrespective of the age of the memory (Moscovitch et al., 2005).

A central problem for addressing these debates with neuropsychology and, more specifically, for determining the relevance of wider brain network abnormalities, is that HPC amnesia is very rare. Consequently, most studies rely upon just a handful of amnesic cases and are thus susceptible to confounds such as impairment severity, measurement noise and individual differences (Lambon Ralph et al., 2011; Lambon Ralph et al., 2002). Larger cohort studies would partially overcome these confounds and capitalize on case heterogeneity to map behavior to brain structure and function.

In order to explore the impact of HPC damage on wider brain networks and determine the impact of these abnormalities upon memory, we studied a uniquely large cohort (n = 38) of patients after autoimmune limbic encephalitis (LE). In its acute phase, this highly consistent clinical syndrome classically causes focal HPC damage, seen on MRI as high T2 signal (Finke et al., 2017; Kotsenas et al., 2014; Loane et al., 2019; Malter et al., 2014). Although most patients respond well to early immunosuppressive therapy (Finke et al., 2017; Irani et al., 2011; Thompson et al., 2018), some subsequently develop HPC atrophy and persistent cognitive impairment, the most prominent aspect of which is anterograde amnesia (Butler et al., 2014; Finke et al., 2017; Irani et al., 2013; Loane et al., 2019; Malter et al., 2014). Autoimmune LE patients are often included in studies of HPC amnesia (Hassabis et al., 2007; Henson et al., 2017; Henson et al., 2016; Maguire et al., 2006). Post-mortem studies demonstrate focal HPC damage (Khan et al., 2009; Park et al., 2007), contrasting with the wider damage typically found following other encephalitides [e.g. herpes simplex encephalitis (Damasio and Van Hoesen, 1985; Gitelman et al., 2001)] or global ischemia/anoxia (Huang and Castillo, 2008), conditions that have also been used as models of human HPC function (for discussion, see Squire and Zola, 1996). Animal models have also shown focal limbic involvement (HPC, amygdala) (Tröscher et al., 2017). Patients’ varying degrees of residual symptom severity offered us the opportunity to examine the relationships between cognition and brain structure and function.

Patients (n = 38) and healthy controls (n = 41) were assessed with a thorough battery of neuropsychological tests of episodic memory (for verbal and visual material) and other cognitive functions, in order to identify the tasks in which patients showed impaired performance. The tests we employed are robust, widely used, well-normed, and largely model-free (see below). Using manual volumetry of MTL regions, as well as whole-brain, voxel-based investigations of structural MRI, we identified regions of gray matter (GM) volume reduction in patients as compared with healthy controls. Using resting-state fMRI (rsfMRI), we also identified resting-state functional abnormalities in patients with respect to both segregation and integration, that is in terms of hemodynamic activity in local regions (resting-state amplitude of low-frequency fluctuations; rsALFF) and functional connectivity between regions (resting-state functional connectivity; rsFC). Finally, we investigated the relationships between brain abnormalities and cognitive deficits. We predicted that: i) HPC damage (documented to occur focally in autoimmune LE) would be accompanied by remote abnormalities in hubs of the extended HPC system, including the HPC-diencephalic-cingulate network (Aggleton, 2014; Bubb et al., 2017); ii) variation in the extent of these network-wide abnormalities across patients would explain variability in memory impairment over and above HPC atrophy; iii) Moreover, on our assumption that HPC damage leads to broader network disruption, we predicted that relationships between HPC atrophy and memory impairment would be mediated by this network disruption. This prediction is in line with recent functional imaging and lesion studies that emphasize the importance of broader networks in episodic memory (Cook et al., 2015; Henson et al., 2016; Inhoff and Ranganath, 2017; Schedlbauer et al., 2014; Watrous et al., 2013); iv) taking these abnormalities into account would enable us to identify which particular aspects of memory impairment are a direct function of HPC atrophy.

Results

Our analysis approach is summarized in Figure 1. We first (1) identified cognitive deficits by comparing the performance of patients and healthy controls on a broad range of neuropsychological tests. We then (2) identified regions in which patients showed structural and (3) functional abnormalities relative to healthy controls. We then (4) examined the relationship between structural/functional brain abnormalities and memory impairment across patients.

Figure 1

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Outline of Results Section.

We first (1) identified cognitive deficits by comparing patients with healthy controls in a broad range of tests of neuropsychological assessment. We identified regions in which patients showed (2) reduced gray matter volumes and (3) resting-state functional connectivity and activity relative to healthy controls; (4) we also identified relationships between structural/functional abnormalities and performance in tests in which patients showed impairment as compared with healthy controls; 'connectome-MVPA': connectome ‘multi-variate pattern analysis’ (Whitfield-Gabrieli and Nieto-Castanon, 2012); MRI: Magnetic Resonance Imaging; MTL: medial temporal lobe; n: number of participants; rsALFF: resting-state amplitude of low-frequency fluctuations; rsFC: resting-state functional connectivity; VBM: voxel-based morphometry.

https://doi.org/10.7554/eLife.46156.002

Neuropsychological assessment

The patients included in this study (n = 38) had been diagnosed with autoimmune LE, and were recruited post-acutely, after reassessment by an experienced neurologist (CRB) prior to study inclusion (see Materials and methods).

Patients did not differ from healthy controls (n = 41; matched for age and M:F ratio) in premorbid intelligence, semantic memory and language, visuomotor and executive function. Their cognitive profile was characterized by highly focal episodic memory impairment. Patients were impaired in both recall and recognition of both verbal and visual material (including scenes), and showed pronounced forgetting of visual material – verbal forgetting was marginally impaired. A clear exception was patients’ preserved face recognition memory. They also showed impaired remote autobiographical memory for both childhood and early adulthood epochs, but preserved remote personal semantic memory. Although they showed higher scores on the depression sub-scale of the Hospital Anxiety and Depression Scale (HADS) (Zigmond and Snaith, 1983), the median score was well below the clinical cut-off, and none of the patients scored within the severe range (mild range: three patients; moderate range: four patients; non-case range: rest of patients and all controls) (Table 1).

Table 1

Neuropsychological profile of autoimmune LE patients (post-acute phase) and healthy controls

https://doi.org/10.7554/eLife.46156.003

Domain		Test	Subtest	Controls		Patients		Controls vs patients			‘Impaired' range
Domain		Test	Subtest	M	IQR	M	IQR	Test	Statistic	p-corr	cut-off score	Patients (n)	Controls (n)
Episodic Memory	Immediate Verbal Recall	WMS-III	Logical Memory I (z)	0.33	1.92	−1.00	1.34	t	6.78	<0.0005	≤ - 1.67	14	0
		WMS-III	Word List I (z)	0.67	1.83	−1.00	1.51	U	178.00	<0.0005	≤ - 1.67	11	0
		D and P	People (z)	−0.33	1.59	−1.33	1.00	U	266.00	<0.0005	≤ - 1.67	15	1
	Delayed Verbal Recall	WMS-III	Logical Memory II (z)	0.67	1.67	−2.00	2.00	U	106.50	<0.0005	≤ - 1.67	23	0
	Delayed Verbal Recall	WMS-III	Word List II (z)	1.33	1.00	−0.67	1.83	U	179.00	<0.0005	≤ - 1.67	3	0
	Verbal Forgetting	D and P	Verbal Forgetting (z)	0.67	1.00	−0.33	1.75	U	415.50	0.0584	≤ - 1.67	8	1
	Verbal Recognition	D and P	Names (z)	0.33	2.00	−1.00	2.00	t	5.16	<0.0005	≤ - 1.67	13	1
		RMT	Words (z)	1.00	1.51	0.00	2.26	U	317.50	<0.0005	≤ - 1.67	9	4
		WMS-III	Word List II Recognition (z)	0.67	1.00	−0.67	2.16	U	266.50	<0.0005	≤ - 1.67	9	0
	Immediate Visual Recall	D and P	Shapes (z)	0.67	1.00	−0.84	2.08	Wt	5.78	<0.0005	≤ - 1.67	13	0
	Immediate Visual Recall	ROCFT	Immediate Recall (z)	1.26	1.93	−0.93	2.58	U	255.50	<0.0005	≤ - 1.67	12	1
	Delayed Visual Recall	ROCFT	Delayed Recall (z)	1.26	1.98	−1.37	3.55	U	258.00	<0.0005	≤ - 1.67	16	1
	Visual Forgetting	D and P	Visual Forgetting (z)	0.33	0.00	0.33	1.83	U	433.00	0.0098	≤ - 1.67	8	1
	Visual Recognition	D and P	Doors (z)	0.67	1.33	−0.67	1.75	t	3.74	0.0072	≤ - 1.67	7	1
		RMT	Scenes (z)	1.00	0.99	−0.35	2.65	U	281.00	0.0025	≤ - 1.67	9	0
		RMT	Faces (z)	0.00	2.33	−0.33	1.66	t	1.29	0.6536	≤ - 1.67	8	6
	Autobiographical Memory	AMI	Childhood (9)	9.00	3.00	5.00	4.00	U	174.50	<0.0005	≤3.00 *	11	1
	Autobiographical Memory		Early Adulthood (9)	9.00	1.50	4.00	4.00	U	123.00	<0.0005	≤3.00 *	14	0
Intelligence, Semantic Memory, and Language	Personal Semantic Memory		Childhood (21)	19.50	3.00	18.00	5.00	U	267.00	0.0835	≤11.00 *	3	0
	Personal Semantic Memory		Early Adulthood (21)	20.50	2.00	19.00	2.50	U	263.00	0.0687	≤14.00 *	4	1
		NART	p-FSIQ (z)	1.44	0.85	1.04	1.05	U	486.00	0.1281	≤ - 1.67	0	0
		WASI/	Vocabulary (z)	1.40	1.25	0.70	1.20	t	3.05	0.0584	≤ - 1.67	0	0
		WASI-II	Similarities (z)	1.05	0.80	0.70	0.85	U	378.00	0.1024	≤ - 1.67	0	0
		GNT	(z)	0.63	0.98	0.15	1.89	U	423.50	0.0683	≤ - 1.67	5	0
		C and CT	(z)	0.34	1.22	0.02	1.22	U	496.50	0.1484	≤ - 1.67	5	0
Executive Function		WMS-III	Digit Span (z)	0.84	1.25	0.33	1.67	t	2.70	0.1024	≤ - 1.67	2	1
Executive Function		DKEFS Trails	Number-Letter Switching (z)	0.67	0.67	0.33	1.00	U	470.00	0.1024	≤ - 1.67	3	1
Visuomotor Function			Visual Scanning (z)	0.67	1.50	0.00	1.34	U	584.00	0.6536	≤ - 1.67	5	4
			Motor Speed (z)	0.67	1.00	0.33	1.34	U	552.00	0.4915	≤ - 1.67	7	5
		ROCFT	copy rank	> 16^th %ile	0.00	> 16^th %ile	0.00	U	619.00	0.4915	≤ 16^th %ile	2	1
		VOSP	Cube Analysis (z)	10.00	1.00	9.00	2.00	U	548.00	0.3116	≤6.00 **	3	0
			Dot Counting (z)	10.00	0.00	10.00	0.00	U	655.50	0.6536	≤8.00 **	1	1
			Position Discrimination (z)	20.00	0.00	20.00	1.00	U	673.00	0.6536	≤18.00 **	4	2
Mood		HADS	Anxiety (21)	4.00	4.00	5.00	5.50	U	420.00	0.0910	≥15.00 ***	3	0
Mood		HADS	Depression (21)	1.00	1.00	3.00	4.50	U	298.00	0.0006	≥15.00 ***	0	0

AMI: Autobiographical Memory Interview; D and P: Doors and People Test; DKEFS: Delis-Kaplan Executive Function System; GNT: Graded Naming Test; HADS: Hospital Anxiety and Depression Scale; IQR: Inter-Quartile Range; M: median; NART: National Adult Reading Test; p-corr: p values are corrected using the Holm-Bonferroni sequential correction for multiple comparisons (n = 35); RMT: Warrington Recognition Memory Tests (words, faces) and Warrington Topographical Memory test (scenes); ROCFT: Rey-Osterrieth Complex Figure Test; t: Student’s t-test; U: Mann-Whitney U; VOSP: Visual Object and Space Perception Battery; WASI/WASI-II Wechsler Abbreviated Scale of Intelligence; WMS-III: Wechsler Memory Scale III; Wt: Welch’s t-test; *,**,***: no standardized scores available for these subtests; *: highest score of ‘definitely abnormal’ range, that is scores at or below which none of the healthy controls scored in Kopelman et al. (1989); **: 5% cut-off score; ***: cut-off score for severe range.

For further correlational analyses, and in order to minimize the contribution of measurement error and maximize the generalizability of our findings, we derived three composite memory scores per participant, in relation to the aforementioned debates in the literature of HPC amnesia. These composite measures comprised scores on individual tests in which patients showed impaired performance at group level compared with controls: i) a composite score for anterograde retrieval, comprising scores on tests of verbal recall, verbal recognition, visual recall, and visual recognition, derived by averaging the corresponding standardized age-scaled scores; ii) a score for anterograde retention (‘forgetting’), which was the visual forgetting score in the Doors and People test (Baddeley et al., 1994); iii) a remote autobiographical memory score, calculated by summing the scores on autobiographical memories for childhood and early adulthood epochs from the Autobiographical Memory Interview (Kopelman et al., 1989) (Supplementary Table 1 in Supplementary file 1).

Structural abnormalities

Volumetry

Consistent with the neuroradiological reports on patients’ acute clinical T2-weighted MRI scans (Table 2) and the fact that acute HPC T2 hyperintensity and oedema are followed by post-acute HPC atrophy in autoimmune LE (Finke et al., 2017; Irani et al., 2013; Loane et al., 2019), volumetric analysis of patients’ post-acute MRIs revealed pronounced bilateral HPC atrophy (left HPC: F = 46.02, p-corr <0.0005; right HPC: F = 63.38, p-corr <0.0005). We also observed right entorhinal (F = 10.76, p-corr = 0.0308) and left thalamic volume reduction (F = 15.41, p-corr = 0.0003; Table 3).

Table 2

Clinical details of autoimmune LE patients (acute phase).

https://doi.org/10.7554/eLife.46156.022

Code	Age (years)	Sex	Antibody type	Acute T2 scan notes
				HPC		Other structures
				R	L	Other structures
1	65.75	M	LGI1	Normal T2 signal and volume; facilitated diffusion	High T2 signal; swelling; normal diffusion	L AMG: high T2 signal
2	69.98	F	VGKCC	Normal T2 signal; mild atrophy; facilitated diffusion	High T2 signal; normal volume; facilitated diffusion	No abnormalities
3	62.23	M	VGKCC	Normal T2 signal and volume; facilitated diffusion	High T2 signal; swelling; normal diffusion	L AMG, L ERC: high T2 signal
4	46.41	M	LGI1	High T2 signal; normal volume; normal diffusion	Normal T2 signal and volume; facilitated diffusion	No abnormalities
5	56.65	M	LGI1	L/R: high T2 signal		No abnormalities
6	58.18	M	LGI1	No abnormalities
7	56.13	M	LGI1	Normal volume and signal	High T2 signal; swelling	L/R AMG: high T2 signal
8	76.54	M	LGI1	High T2 signal; normal volume and diffusion	High T2 signal; normal volume; facilitated diffusion	No abnormalities
9	54.94	M	LGI1	High T2 signal; swelling; normal diffusion	High T2 signal; swelling; normal diffusion	No abnormalities
10	44.81	M	LGI1	L/R: high T2 signal; swelling		No abnormalities
11	45.77	M	LGI1	High T2 signal; normal volume	High T2 signal; normal volume	No abnormalities
12	46.06	M	LGI1/Caspr2	High T2 signal; atrophy	Normal T2 signal and volume	No abnormalities
13	35.75	M	LGI1/Caspr2	L/R: normal T2 signal; mild atrophy; normal diffusion		No abnormalities
14	72.08	M	LGI1	High T2 signal; mild atrophy; facilitated diffusion	Normal T2 signal; atrophy; facilitated diffusion	No abnormalities
15	52.28	M	LGI1	High T2 signal; normal volume; facilitated diffusion	Normal T2 signal and volume; facilitated diffusion	No abnormalities
16	52.48	M	LGI1/Caspr2	High T2 signal; swelling; facilitated diffusion	Normal T2 signal and volume; facilitated diffusion	No abnormalities
17	51.62	M	VGKCC	High T2 signal; swelling	Normal T2 signal and volume	No abnormalities
18	75.18	M	LGI1	L/R: high T2 signal; swelling; normal diffusion		L/R AMG: high T2 signal; swelling
19	78.73	M	LG1/Caspr2	High T2 signal; mild atrophy; normal diffusion	High T2 signal; normal volume; normal diffusion	No abnormalities
20	53.75	F	LGI1	L/R: high T2 signal; normal volume and diffusion		No abnormalities
21	73.68	F	VGKCC	L/R: high T2 signal; swelling; facilitated diffusion		No abnormalities
22	63.59	M	LGI1	L/R: high T2 signal; normal volume and diffusion		No abnormalities
23	60.35	M	VGKCC	No abnormalities
24	54.30	M	VGKCC	L/R: high T2 signal; atrophy		L/R AMG: high T2 signal; atrophy
25	52.70	M	seronegative	L/R: high T2 signal		No abnormalities
26	47.43	F	seronegative	No abnormalities
27	58.60	M	seronegative	L/R: high T2 signal		No abnormalities
28	25.42	M	Anti-Ma2	L/R: high T2 signal and swelling		No abnormalities
29	45.77	F	seronegative	L/R: high T2 signal		No abnormalities
30	16.64	F	GAD	No abnormalities
31	71.35	M	seronegative	L/R: high T2 signal; atrophy		No abnormalities
32	60.44	M	VGKCC	L/R: atrophy		PHC atrophy
33	53.48	M	seronegative	L/R: atrophy		No abnormalities
34	64.87	F	seronegative	L/R: atrophy		No abnormalities
35	47.32	F	seronegative	L/R: high T2 signal		R AMG: high T2 signal; swelling
36	61.88	F	seronegative	L/R: high T2 signal; atrophy		No abnormalities
37	71.90	F	seronegative	L/R: high T2 signal (especially R)		No abnormalities
38	34.49	F	GAD	L/R: high T2 signal		No abnormalities

Age: age at symptom onset (years); AMG: Amygdala; Caspr2: anti-contactin-associated protein-like 2; ERC: entorhinal cortex; F = female; GAD: anti-glutamic acid decarboxylase autoantibody; HPC: hippocampus; L: left hemisphere; LGI1: anti-leucine-rich glioma-inactivated1; M = male; PHC: parahippocampal cortex; R: right hemisphere; VGKCC: anti-voltage-gated potassium channel complex. The clinical details of patients 1–24 have also been presented in Loane et al. (2019).

Table 3

Volumetry of MTL and subcortical structures in autoimmune LE patients.

https://doi.org/10.7554/eLife.46156.004

Structure	Controls		Patients		Mean % reduction	F	Partial η²	p-corr
Structure	Mean (mm³)	SD (mm³)	Mean (mm³)	SD (mm³)	Mean % reduction	F	Partial η²	p-corr
R HPC	3648.99	459.59	2733.87	751.09	−25.08	63.38	0.390	<0.0005
L HPC	3439.48	431.91	2671.18	710.65	−22.34	46.02	0.317	<0.0005
R ERC	1602.69	324.07	1254.97	404.70	−21.70	10.76	0.119	0.0308
L ERC	1508.83	326.07	1200.18	432.07	−20.46	9.48	0.106	0.0534
R Thalamus	7407.51	762.15	7072.79	845.09	−4.52	8.23	0.077	0.0900
L Thalamus	7633.09	788.11	7194.76	797.73	−5.74	15.41	0.135	0.0034
R PRC	1791.42	378.97	1561.61	403.86	−12.83	7.11	0.082	0.1575
L PRC	1812.50	523.31	1601.39	478.78	−11.65	2.88	0.035	>0.9999
R PHC	1900.73	423.85	1665.68	331.28	−12.37	6.36	0.074	0.2240
L PHC	2016.52	435.92	1851.53	445.52	−8.18	1.53	0.019	>0.9999
R AMG	1395.67	267.38	1313.76	406.51	−5.87	5.15	0.060	0.3899
L AMG	1321.96	217.40	1268.47	384.22	−4.05	4.43	0.053	0.5320
R Nacc	339.42	107.77	318.53	109.69	−6.16	0.25	0.003	>0.9999
L Nacc	433.79	127.41	381.89	153.66	−11.96	3.70	0.036	0.7410
R TPC	4558.60	846.26	4520.16	1026.08	−0.84	0.12	0.002	>0.9999
L TPC	4331.40	742.35	4512.95	795.76	4.19	1.90	0.023	>0.9999
R Putamen	4332.54	548.04	4157.00	630.21	−4.05	0.85	0.008	>0.9999
L Putamen	4382.39	705.40	4151.89	662.93	−5.26	1.67	0.017	>0.9999
R Caudate	3403.96	440.42	3369.16	453.63	−1.02	0.06	0.001	>0.9999
L Caudate	3211.40	438.40	3134.26	497.42	−2.40	0.12	0.001	>0.9999
R Pallidum	1705.61	260.63	1630.00	251.08	−4.43	0.90	0.009	>0.9999
L Pallidum	1720.40	297.86	1661.13	308.18	−3.45	0.41	0.004	>0.9999
brainstem	22119.06	2258.25	21931.42	2647.93	−0.85	0.23	0.002	>0.9999

Volumetry of manually and automatically delineated MTL and other subcortical structures of all patients (n = 38). Volumes for each structure are compared between patients and controls, using age, sex, TIV, and scan source (MAP, OPTIMA) as between-subjects covariates in a series of univariate ANCOVAs; AMG: amygdala; ANCOVA: analysis of covariance; ; ERC: entorhinal cortex; HPC: hippocampus; L: left hemisphere; MAP: Memory and Amnesia Project; MTL: medial temporal lobe; Nacc: nucleus accumbens; OPTIMA: Oxford Project To Investigate Memory and Aging; p-corr: p values are adjusted with the Holm-Bonferroni sequential correction method for multiple comparisons (n = 23); PHC: parahippocampal cortex; PRC: perirhinal cortex; R: right hemisphere; SD: standard deviation; TIV: total intracranial volume; TPC: temporopolar cortex.

Whole-brain VBM: GM volume

Strongly consistent with the volumetric findings above, a whole-brain VBM contrast disclosed reduced GM volume in patients’ left and right HPC, as well as in the anterior/mediodorsal, and right dorsolateral regions of the thalamus (Figure 2a; Table 4).

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Reduction in GM volume, rsALFF and rsFC in autoimmune LE patients.

(a) A whole-brain VBM on GM volume (contrast: controls > patients) showed volume reduction in patients’ HPC bilaterally, as well as in mediodorsal-anterior and right dorsolateral thalamic regions (Table 4); clusters survive FWE peak-level correction (p<0.05) over p<0.001 unc; color bar indicates t values; b-c: Reduced rsALFF in patients in (b) the posterior cingulate (kE = 89, p-FWE = 0.033; peak voxel coordinates: −4,–36, 28) and (c) precuneus (kE = 137; p-FWE = 0.003; peak voxel coordinates: 4,–60, 26); d-j: reduced rsFC in patients; d: a whole-brain MVPA (omnibus F) showed abnormal rsFC for patients in a cluster in the right HPC (kE = 178, p-FWE = 0.001; peak voxel coordinates: 28,–16,−20; color bar indicates F values); e-g: reduced rsFC of the right HPC (whole-brain seed-to-voxel analysis; seed: right HPC, anatomically delineated in native space, unsmoothed timeseries; contrast: controls > patients); e: medial prefrontal cortex (kE = 1152, p-FWE <0.0001, peak voxel coordinates: 4, 56, 2); f: posteromedial cortex (posterior cingulate, retrosplenial cortex, precuneus; kE = 986, p-FWE <0.0001, peak voxel coordinates: 6,–50, 8); g: left HPC (kE = 393, p-FWE <0.0001, peak voxel coordinates: −12,–36, 2). All rsFC and rsALFF clusters survive FWE correction (p<0.05) for cluster size over an individual voxel threshold of p<0.001; FWE: family-wise error; HPC: hippocampus; kE: cluster size (number of voxels); rsALFF: Resting-state amplitude of low frequency fluctuations; rsFC: Resting-state functional connectivity; VBM: voxel-based morphometry.

https://doi.org/10.7554/eLife.46156.005

Table 4

GM volume reduction in autoimmune LE patients (whole-brain VBM).

https://doi.org/10.7554/eLife.46156.008

kE	Peak					Center of mass			Structure
kE	p-FWE	T	X	Y	Z	X	Y	Z	Structure
2574	<0.0005	7.53	28	−17	−20	28	−16	−18	R HPC
910	<0.0005	6.81	−29	−12	−18	−27	−15	−19	L HPC
113	0.002	6.18	19	−28	5	18	−26	6	R lateral thalamus
414	0.006	5.85	-1	−16	0	3	−12	7	anterior/mediodorsal thalamus

Contrast: controls > patients; covariates: age, sex, scan source (MAP, OPTIMA), and TIV. Clusters are FWE-corrected at peak-voxel level (p<0.05) over an individual voxel threshold of p<0.001 (unc.); voxel size: 1 mm³ isotropic; spatial smoothing kernel: 4 mm FWHM; FWHM: Full-width at half-maximum; HPC: hippocampus; kE: cluster size (number of voxels; minimum cluster size: 50 voxels); L: Left hemisphere; mm: millimeter; MAP: Memory and Amnesia Project; OPTIMA: Oxford Project To Investigate Memory and Aging; R: Right hemisphere; TIV: total intracranial volume; x, y, z: coordinates in mm.

Functional abnormalities

We also investigated resting-state functional abnormalities in patients with respect to hemodynamic activity in local regions (resting-state amplitude of low-frequency fluctuations; rsALFF) and functional connectivity between regions, in the form of resting-state functional connectivity (rsFC) (see Materials and methods section), across the whole brain and in a data-driven fashion.

Resting-state hemodynamic activity

In order to identify specific brain regions that show abnormal hemodynamic activity at rest in patients, similar to resting-state CBF and glucose metabolic rate in PET studies, we analyzed rsALFF, that is the intensity of slow spontaneous fluctuations of hemodynamic activity at rest across the whole brain (Zang et al., 2007).

As compared with healthy controls, patients showed reduced rsALFF in the posterior cingulate cortex (PCC) and the precuneus (Figure 2b–c). Consistent with the reciprocal connectivity of thalamic nuclei with both the HPC and the cingulate cortex (Aggleton, 2014; Aggleton et al., 2010; Bubb et al., 2017) and our hypothesis that HPC atrophy is followed by structural and functional abnormalities in interconnected areas within the HPC-diencephalic-cingulate networks, we observed that the effect of average HPC volume reduction on the rsALFF of the PCC was fully mediated by average thalamic volume reduction (Figure 2—figure supplement 1; Figure 2—figure supplements 1—source data 1).

Resting-state functional connectivity

Voxel-to-Voxel rsFC: connectome-MVPA

Capitalizing on the size of our patient cohort, we conducted a data-driven, whole-brain, principal components-based analysis, (‘connectome-MVPA’), implemented in the Conn toolbox (Whitfield-Gabrieli and Nieto-Castanon, 2012). This method is used to identify the voxel clusters in which healthy controls and patient groups differ significantly with respect to their rsFC with the rest of the brain, instead of selecting seed/target regions or networks in an a priori fashion.

This analysis showed group differences in the whole-brain rsFC of a right HPC cluster (Figure 2d).

Seed-to-Voxel rsFC

In order to identify the specific brain regions showing reduced rsFC with the right HPC in patients, the right HPC was then selected for a whole-brain seed-to-voxel rsFC analysis (Biswal et al., 1995; Margulies et al., 2007). The spatially unsmoothed timeseries data were extracted from participants’ manually delineated right HPC in native space, ensuring that rsFC differences were not an artefact of insufficient co-registration of the atrophic HPC. This seed region showed reduced rsFC with clusters in the medial prefrontal and posteromedial (PCC, retrosplenial, and precuneus) cortices, and the left HPC (Figure 2e–g). We also wanted to ensure that the left HPC cluster that showed reduced rsFC with the right HPC was not a result of suboptimal co-registration of the functional images with the atrophic HPC. For further correlational analyses (section below), we thus used the mean rsFC values between the right and left HPC in native space (unsmoothed time series) instead of the left HPC cluster. Patients’ reduced inter-HPC rsFC (of all 13 structural and functional abnormalities) negatively correlated, across patients, with the delay between the onset of their symptoms and the time they underwent our research MRI (Supplementary Table 2 in Supplementary file 1).

Structure/Function-Behavior Correlations

Having identified the core brain abnormalities in our patient group, we investigated the contributions of these to explaining memory impairment.

We first applied a stringent correction for multiple testing for the total number of correlations conducted (n = 39) between the brain abnormalities identified (n = 13; right/left HPC volumes and right entorhinal cortical volumes, based on manual delineation; left thalamic volumes, based on automated segmentation; anterior-mediodorsal and right dorsolateral thalamic volumes and right/left HPC volumes, expressed by the VBM clusters above; reduced rsALFF in the precuneus and the PCC; reduced rsFC between the right HPC and the left HPC, the medial prefrontal cortex, and the precuneus) and the three composite memory scores (n = 3; anterograde retrieval; anterograde retention; remote autobiographical memory). Three correlations survived correction for multiple tests: i) Anterograde retrieval scores correlated across patients with their reduced rsALFF in the PCC (r = 0.551; p-corr = 0.024); ii) Anterograde retention (i.e. ‘forgetting’) scores correlated with patients’ reduced right HPC volume (VBM cluster; rho = 0.556, p-corr = 0.024; manually delineated right HPC volume: rho = 0.508, p-corr = 0.079); iii) Remote autobiographical memory scores correlated with patients’ reduced volume in the left thalamus (r = 0.558; p-corr = 0.041; rest of ps, p-corr ≥0.105; Supplementary Table 3 in Supplementary file 1).

Given the striking lack of correlations with HPC volume, we addressed the possibility of false negatives in our original approach by iterating the correlational analyses above after introducing three amendments: i) we fragmented the anterograde retrieval composite score into four composite scores (visual/verbal recall/recognition), taking into account the possibility of different relationships of recall vs. recognition memory scores with brain abnormalities; ii) we applied a more lenient correction for the number of structural/functional abnormalities (n = 13), separately for each composite score examined (n = 6; visual/verbal recall/recognition, remote autobiographical memory, visual forgetting; Supplementary Table 4 in Supplementary file 1); iii) in a post-hoc fashion, we examined the relationship of these memory scores with the manually delineated anterior vs. posterior HPC portions at uncorrected levels (Supplementary Table 5 in Supplementary file 1).

Anterograde memory: Verbal recognition

Verbal recognition scores correlated with patients’ reduced inter-HPC rsFC (r = 0.498, p-corr = 0.039; rest of ps, p-corr ≥0.18; Figure 3b; Figure 3—source data 1). Out of the HPC volumes, only the volume expressed by the left HPC VBM cluster correlated with verbal recognition scores at uncorrected levels (r = 0.360, p-unc = 0.029; Figure 3a; rest of rs, 0.281 ≥ r ≥ 0.276; rest of ps, 0.098 ≥ p unc ≥ 0.092). None of the HPC volumes examined (left/right HPC VBM clusters; manually delineated left/right HPC) correlated with inter-HPC rsFC across patients (all rs, |r| ≤ 0.135; all ps, p-unc ≥ 0.441). We thus entered mean inter-HPC rsFC and one of those four HPC volume measures as independent variables in four separate multiple step-wise linear regressions, with verbal recognition scores as the dependent variable. In all four analyses, the regression terminated in a single step, with inter-HPC rsFC as the only predictor of patients’ performance (R² = 0.25; β(z) = 0.50; F = 10.53, p = 0.003).

Figure 3

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Verbal Recognition Memory: Structural/Functional correlates.

( a) GM volume expressed by the left HPC VBM cluster correlated across patients with their verbal recognition composite memory scores only at uncorrected levels (r = 0.36, p-unc = 0.029); (b) mean inter-HPC rsFC correlated across patients with their verbal recognition composite memory scores and survived correction for multiple (structural/functional brain abnormalities examined: n = 13) testing (r = 0.50, p-corr = 0.039); GM: gray matter; HPC: hippocampus; L, R: left, right (hemisphere); MAP: Memory and Amnesia Project; OPTIMA: Oxford Project To Investigate Memory and Aging; p: significance values are presented at uncorrected levels; rho: Spearmann’s rank correlation coefficient; rsFC: resting-state functional connectivity; VBM: voxel-based morphometry; z: memory scores are averaged age-scaled and standardized scores of participants’ performance in the subtests of interest; z-res: GM volume from VBM clusters is residualized against age, sex, scan source (MAP, OPTIMA) and TIV across participants; mean rsFC is residualized across participants against age and sex.

https://doi.org/10.7554/eLife.46156.009

Figure 3—source data 1 This spreadsheet contains the mean GM volume of the left HPC VBM cluster and the mean inter-HPC rsFC (z-res) and the verbal recognition memory composite scores (z) of healthy controls and patients that are plotted in Figure 3; GM: gray matter; HPC: hippocampus; MAP: Memory and Amnesia Project; OPTIMA: Oxford Project To Investigate Memory and Aging; rsFC: resting-state functional connectivity; VBM: voxel-based morphometry; z: memory scores are averaged age-scaled and standardized scores of participants’ performance in the subtests of interest; z-res: GM volumes from VBM clusters are residualized against age, sex, scan source (MAP, OPTIMA), and TIV across participants; mean rsFC is residualized against age and sex across participants. These data can be opened with Microsoft Excel or with open-source alternatives such as OpenOffice.: https://doi.org/10.7554/eLife.46156.010
Download elife-46156-fig3-data1-v2.xlsx

Anterograde memory: Visual recognition

Visual recognition scores correlated with patients’ reduced rsALFF in the PCC (r = 0.543, p-corr = 0.014), and only marginally with the volume of the left HPC VBM cluster (r = 0.449, p-corr = 0.072; rest of ps, p-corr ≥0.110). The volume of the left HPC VBM cluster correlated with the rsALFF in the PCC across patients (r = 0.449, p=0.007).

Patients’ reduced rsALFF in the PCC correlated with visual recognition memory scores over and above the correlative reduction in the left HPC volume (VBM cluster) (partial correlation: r = 0.432, p=0.013). Since left HPC volumes correlated across patients with their reduced rsALFF in the PCC, and since both these abnormalities correlated with patients’ impaired visual recognition memory, we conducted a mediation analysis to test our hypothesis that the effects of HPC damage trigger abnormalities within the extended HPC system, and that these underlie memory impairment (see Materials and methods section). Indeed, the effects of left HPC volume reduction on visual recognition scores were fully mediated by the reduction in PCC rsALFF (direct effect: β = 0.24, p=0.183; indirect effect: β = 0.21, 95% CI: 0.03, 0.41) (Figure 4; Figure 4—source data 1). In other words, controlling for the mediator variable (rsALFF in the PCC) reduced the variance of the dependent variable (scores of visual recognition memory) explained by the independent variable (volume expressed by the left HPC VBM cluster), to the extent that the relationship between the independent and dependent variables became non-significant.

Figure 4

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Visual Recognition Memory: Structural/Functional correlates.

(a) mean GM volume of the left HPC cluster correlated with the mean rsALFF of the PCC cluster across patients; (b) visual recognition memory scores correlated across patients with their mean rsALFF in the PCC cluster, surviving correction for multiple testing for the 13 structural/functional abnormalities examined (r = 0.54, p-corr = 0.014); the mediation analysis demonstrates that this effect held over and above the correlation of PCC rsALFF with the mean GM volume of the left HPC cluster; (c) mean GM volume of the left HPC cluster correlated with visual recognition memory scores across patients, but did not survive correction for multiple testing (r = 0.45, p-corr = 0.072); however, the mediation analysis demonstrated that this relationship did not hold over and above the correlation of the mean GM volume of the left HPC cluster with the mean PCC rsALFF; there was only an indirect effect of reduced HPC GM volume on visual recognition memory (within parenthesis: 95% confidence intervals); GM: gray matter; HPC: hippocampus; L: left (hemisphere); MAP: Memory and Amnesia Project; OPTIMA: Oxford Project To Investigate Memory and Aging; p: significance values are presented at uncorrected levels; PCC: posterior cingulate cortex; rsALFF: resting-state amplitude of low frequency fluctuations; TIV: total intracranial volume; VBM: voxel-based morphometry; z: memory scores are averaged age-scaled and standardized scores of participants’ performance in the subtests of interest; z-res: GM volumes from VBM clusters are residualized against age, sex, scan source (MAP, OPTIMA), and TIV across participants; mean rsALFF values are residualized across participants against age and sex.

https://doi.org/10.7554/eLife.46156.011

Figure 4—source data 1 This spreadsheet contains the mean GM volume of the left HPC VBM cluster and the mean rsALFF in the PCC (z-res) and the visual recognition memory composite scores (z) of healthy controls and patients that are plotted in Figure 4; GM: gray matter; HPC: hippocampus; MAP: Memory and Amnesia Project; OPTIMA: Oxford Project To Investigate Memory and Aging; PCC: posterior cingulate cortex; rsALFF: resting-state amplitude of low-frequency fluctuations; VBM: voxel-based morphometry; z: memory scores are averaged age-scaled and standardized scores of participants’ performance in the subtests of interest; z-res: GM volumes from VBM clusters are residualized against age, sex, scan source (MAP, OPTIMA), and TIV across participants; mean rsALFF is residualized against age and sex across participants. These data can be opened with Microsoft Excel or with open-source alternatives such as OpenOffice.: https://doi.org/10.7554/eLife.46156.012
Download elife-46156-fig4-data1-v2.xlsx

Anterograde memory: Verbal recall

Verbal recall memory composite scores correlated across patients with their reduced PCC rsALFF (r = 0.582, p-corr = 0.004) and with the volume of the left HPC VBM cluster (r = 0.495, p-corr = 0.024; rest of ps, p-corr ≥0.290). Patients’ reduced rsALFF in the PCC correlated with verbal recall scores over and above the left HPC volume reduction (VBM cluster) (partial correlation: r = 0.474, p=0.005). Consistent with our hypothesis, the effects of HPC atrophy on verbal recall were fully mediated by the reduction in PCC rsALFF (direct effect: β = 0.16, p=0.124; indirect effect: β = 0.14, 95% CI: 0.03, 0.29; Figure 5; Figure 5—source data 1).

Figure 5

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Verbal Recall Memory: Structural/Functional correlates.

(a) mean GM volume of the left HPC cluster correlated with the mean rsALFF of the PCC cluster across patients; (b) verbal recall memory scores correlated across patients with their mean rsALFF in the PCC cluster, surviving corrections for multiple testing across the 13 structural/functional abnormalities examined (r = 0.582, p-corr = 0.004); the mediation analysis demonstrates that this effect held over and above the correlation of PCC rsALFF with the mean GM volume of the left HPC cluster; (c) mean GM volume of the left HPC cluster correlated with verbal recall memory scores across patients (r = 0.495, p-corr = 0.024); however, the mediation analysis demonstrated that this relationship did not hold over and above the correlation of the mean GM volume of the left HPC cluster with the mean PCC rsALFF; there was only an indirect effect of reduced HPC GM volume on verbal recall memory (within parenthesis: 95% confidence intervals); GM: gray matter; HPC: hippocampus; L: left (hemisphere); MAP: Memory and Amnesia Project; OPTIMA: Oxford Project To Investigate Memory and Aging; PCC: posterior cingulate cortex; rsALFF: resting-state amplitude of low frequency fluctuations; TIV: total intracranial volume; VBM: voxel-based morphometry; z: memory scores are averaged age-scaled and standardized scores of participants’ performance in the subtests of interest; z-res: GM volumes from VBM clusters are residualized against age, sex, scan source (MAP, OPTIMA), and TIV across participants; mean rsALFF values are residualized across participants against age and sex.

https://doi.org/10.7554/eLife.46156.013

Figure 5—source data 1 This spreadsheet contains the mean GM volume of the left HPC VBM cluster and the mean rsALFF in the PCC (z-res) and the verbal recall memory composite scores (z) of healthy controls and patients that are plotted in Figure 5; GM: gray matter; HPC: hippocampus; MAP: Memory and Amnesia Project; OPTIMA: Oxford Project To Investigate Memory and Aging; PCC: posterior cingulate cortex; rsALFF: resting-state amplitude of low-frequency fluctuations; VBM: voxel-based morphometry; z: memory scores are averaged age-scaled and standardized scores of participants’ performance in the subtests of interest; z-res: GM volumes from VBM clusters are residualized against age, sex, scan source (MAP, OPTIMA), and TIV across participants; mean rsALFF is residualized against age and sex across participants. These data can be opened with Microsoft Excel or with open-source alternatives such as OpenOffice.: https://doi.org/10.7554/eLife.46156.014
Download elife-46156-fig5-data1-v2.xlsx

Anterograde memory: Visual recall

Visual recall scores correlated at uncorrected levels with PCC rsALFF (rho = 0.446, p-unc = 0.008; p-corr = 0.107), and the left HPC volume (VBM cluster) (rho = 0.370, p-unc = 0.026; p-corr = 0.312; rest of ps, p-corr ≥0.594). Across patients, the reduced rsALFF in the PCC marginally correlated with visual recall memory scores over and above their reduced left HPC volume (VBM cluster) (partial correlation: rho = 0.33, p=0.058). The effects of HPC atrophy on visual recall were fully mediated by the reduction in PCC rsALFF (direct effect: β = 0.23, p=0.272; indirect effect: β = 0.19, 95% CI:0.01,0.46) (Figure 6; Figure 6—source data 1).

Figure 6

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Visual Recall Memory: Structural/Functional correlates.

(a) mean GM volume of the left HPC cluster correlated with the mean rsALFF of the PCC cluster across patients; (b) visual recall memory scores correlated at uncorrected levels across patients with their mean rsALFF in the PCC cluster; the mediation analysis demonstrates that this effect held over and above the correlation of PCC rsALFF with the mean GM volume of the left HPC cluster; (c) mean GM volume of the left HPC cluster correlated with visual recall memory scores across patients at uncorrected levels; however, the mediation analysis demonstrated that this relationship did not hold over and above the correlation of the mean GM volume of the left HPC cluster with the mean PCC rsALFF; there was only an indirect effect of reduced HPC GM volume on visual recall memory (within parenthesis: 95% confidence intervals); GM: gray matter; HPC: hippocampus; L: left (hemisphere); MAP: Memory and Amnesia Project; OPTIMA: Oxford Project To Investigate Memory and Aging; p: significance values are presented at uncorrected levels; PCC: posterior cingulate cortex; rsALFF: resting-state amplitude of low frequency fluctuations; TIV: total intracranial volume; VBM: voxel-based morphometry; z: memory scores are averaged age-scaled and standardized scores of participants’ performance in the subtests of interest; z-res: GM volumes from VBM clusters are residualized against age, sex, scan source (MAP, OPTIMA), and TIV across participants; mean rsALFF values are residualized across participants against age and sex.

https://doi.org/10.7554/eLife.46156.015

Figure 6—source data 1 This spreadsheet contains the mean. GM volume of the left HPC VBM cluster and the mean rsALFF in the PCC (z-res) and the visual recall memory composite scores (z) of healthy controls and patients that are plotted in Figure 6; GM: gray matter; HPC: hippocampus; MAP: Memory and Amnesia Project; OPTIMA: Oxford Project To Investigate Memory and Aging; PCC: posterior cingulate cortex; rsALFF: resting-state amplitude of low-frequency fluctuations; VBM: voxel-based morphometry; z: memory scores are averaged age-scaled and standardized scores of participants’ performance in the subtests of interest; z-res: GM volumes from VBM clusters are residualized against age, sex, scan source (MAP, OPTIMA), and TIV across participants; mean rsALFF is residualized against age and sex across participants. These data can be opened with Microsoft Excel or with open-source alternatives such as OpenOffice.: https://doi.org/10.7554/eLife.46156.016
Download elife-46156-fig6-data1-v2.xlsx

Anterograde memory: Retention (Forgetting)

Anterograde retention (visual forgetting) scores correlated only with volume reduction in the right HPC VBM cluster (rho = 0.556, p-corr = 0.008) and in the manually delineated right HPC (rho = 0.508, p-corr = 0.026; an alternative analysis, comparing patients that scored at ceiling with the rest of the patient group, is reported in Supplementary Table 6 in Supplementary file 1, disclosing the same relationships). No extra-HPC abnormalities correlated significantly with visual forgetting (rest of ps, p-corr ≥0.171). Nevertheless, at uncorrected levels, patients’ reduced rsALFF in the PCC correlated with their visual forgetting scores (rho = 0.399, p-unc = 0.024), as well as with right HPC volume (r = 0.517, p=0.001). Right HPC volume correlated with visual forgetting scores over and above rsALFF in the PCC (partial correlation: rho = 0.460, p=0.009). The strong relationship of visual forgetting scores with right HPC volumes across patients was further demonstrated by a mediation analysis, whereby the effect of right HPC volume reduction on visual forgetting scores remained unmediated by the correlative reduction in PCC rsALFF across patients (direct effect: β = 0.43, p=0.036; indirect effect: β = 0.10, 95% CI: −0.11,0.32; Figure 7; Figure 7—source data 1).

Figure 7

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Forgetting: Structural/Functional correlates.

(a) mean GM volume of the right HPC cluster correlated with the mean rsALFF of the PCC cluster across patients; (b) visual forgetting scores correlated across patients with their mean rsALFF in the PCC cluster, without, however, surviving correction for multiple tests (rho = 0.40, p-corr = 0.216); the mediation analysis demonstrates that this effect did not hold when the correlation of PCC rsALFF with the mean GM volume of the right HPC cluster was accounted for; (c) mean GM volume of the right HPC cluster correlated with visual forgetting scores across patients, surviving correction across the 13 structural/functional abnormalities examined (rho = 0.56, p-corr = 0.008); the mediation analysis demonstrated that this relationship held over and above the correlation of the mean GM volume of the right HPC cluster with the mean PCC rsALFF; there was thus a direct effect of reduced HPC GM volume on visual forgetting (within parenthesis: 95% confidence intervals); D and P: Doors and People (Baddeley et al., 1994); GM: gray matter; HPC: hippocampus; MAP: Memory and Amnesia Project; OPTIMA: Oxford Project To Investigate Memory and Aging; p: significance values are presented at uncorrected levels; PCC: posterior cingulate cortex; R: right (hemisphere); rsALFF: resting-state amplitude of low frequency fluctuations; TIV: total intracranial volume; VBM: voxel-based morphometry; z: age-scaled and standardized scores on Visual Forgetting (D and P); z-res: GM volumes from VBM clusters are residualized against age, sex, scan source (MAP, OPTIMA), and TIV across participants; mean rsALFF values are residualized across participants against age and sex.

https://doi.org/10.7554/eLife.46156.017

Figure 7—source data 1 This spreadsheet contains the mean. GM volume of the right HPC VBM cluster and the mean rsALFF in the PCC (z-res) and the visual forgetting scores (z) of healthy controls and patients that are plotted in Figure 7; GM: gray matter; HPC: hippocampus; MAP: Memory and Amnesia Project; OPTIMA: Oxford Project To Investigate Memory and Aging; PCC: posterior cingulate cortex; rsALFF: resting-state amplitude of low-frequency fluctuations; VBM: voxel-based morphometry; z: age-scaled and standardized scores for visual forgetting [Doors and People (Baddeley et al., 1994)]; z-res: GM volumes from VBM clusters are residualized against age, sex, scan source (MAP, OPTIMA), and TIV across participants; mean rsALFF is residualized against age and sex across participants. These data can be opened with Microsoft Excel or with open-source alternatives such as OpenOffice.: https://doi.org/10.7554/eLife.46156.018
Download elife-46156-fig7-data1-v2.xlsx

Of the three composite memory scores, anterograde retention correlated with scores for depression (HADS) across patients (rho = −0.425, p-corr = 0.045; rest of ps, p-corr >0.248). We thus also examined the relationship among scores for depression, anterograde retention and HPC atrophy, with which anterograde retention scores strongly correlated. No correlation of scores for depression with HPC volumes (VBM clusters, manually delineated volumes) reached significance even at uncorrected levels (all rhos, |rho| ≤ 0.25; all ps, p-unc ≥0.158), and right HPC volumes correlated with visual forgetting scores over and above depression scores (right HPC VBM cluster: rho = 0.564, p=0.001; manually delineated right HPC: rho = 0.530, p=0.002).

Remote autobiographical memory

Remote autobiographical memory scores correlated across patients with their reduced left thalamic volume (r = 0.558, p-corr = 0.015), and only marginally with the volume expressed by the left HPC VBM cluster (r = 0.467, p-corr = 0.096). The volumes of the left thalamus and the left HPC VBM cluster correlated across patients (r = 0.47, p=0.003). Patients’ reduced left thalamic volume correlated with their remote autobiographical memory scores over and above their reduced left HPC volume (VBM cluster) (partial correlation: r = 0.422, p=0.020). Moreover, the effects of left HPC atrophy on remote autobiographical memory were fully mediated by volume reduction in the left thalamus (direct effect: β = 0.94, p=0.165; indirect effect: β = 0.82, 95% CI: 0.20, 1.89; Figure 8; Figure 8—source data 1).

Figure 8

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Remote Autobiographical Memory: Structural/Functional correlates.

(a) GM volume of the left HPC VBM cluster correlated with the left thalamic volume across patients; (b) remote autobiographical memory (AMI) scores correlated across patients with the volume of the left thalamus, surviving correction across the 13 structural/functional abnormalities examined (r = 0.558, p-corr = 0.015); the mediation analysis demonstrates that this effect held when the correlation of thalamic volume with the volume expressed by the left HPC VBM cluster was accounted for; (c) the volume expressed by the left HPC VBM cluster correlated with remote autobiographical memory scores, albeit this correlation did not survive correction for multiple testing (r = 0.467, p-corr = 0.096); the mediation analysis demonstrated that this relationship did not hold over and above the correlation of the left thalamic volume with the HPC clusters; there was thus no direct effect of reduced HPC GM volume on remote autobiographical memory (within parenthesis: 95% confidence intervals); ‘18’: remote autobiographical memory scores are the sums of the AMI scores for autobiographical memories for childhood and early adulthood (max = 18); AMI: Autobiographical Memory Interview; GM: gray matter; HPC: hippocampus; MAP: Memory and Amnesia Project; OPTIMA: Oxford Project To Investigate Memory and Aging; p: significance values are presented at uncorrected levels; TIV: total intracranial volume; VBM: voxel-based morphometry; z-res: volumes are residualized against age, sex, scan source (MAP, OPTIMA) and TIV across participants.

https://doi.org/10.7554/eLife.46156.019

Figure 8—source data 1 This spreadsheet contains the mean GM volume of the left HPC VBM cluster and the volume of the automatically delineated left thalamus (z-res) and the remote autobiographical memory scores [max = 18; Autobiographical Memory Interview; (Kopelman et al., 1989)] of healthy controls and patients (over the age of 50 at the time of assessment; See Materials and methods section) that are plotted in Figure 8; GM: gray matter; HPC: hippocampus; MAP: Memory and Amnesia Project; OPTIMA: Oxford Project To Investigate Memory and Aging; VBM: voxel-based morphometry; z-res: volumes are residualized against age, sex, scan source (MAP, OPTIMA), and TIV across participants. These data can be opened with Microsoft Excel or with open-source alternatives such as OpenOffice.: https://doi.org/10.7554/eLife.46156.020
Download elife-46156-fig8-data1-v2.xlsx

Table 5 summarizes the relationships identified between memory impairment and structural and functional abnormalities in patients, as related to the effects of HPC atrophy. We did not identify any additional relationships when examining the volumes of manually delineated HPC portions (left/right anterior/posterior HPC; Supplementary Table 5 in Supplementary file 1). Moreover, these relationships were not driven by the subset of patients (n = 7) who had not received immunosuppressive therapy (Supplementary Tables 7-8 in Supplementary file 1).

Table 5

Summary of relationships between memory impairment and structural/functional abnormalities across patients.

https://doi.org/10.7554/eLife.46156.021

Memory composite score		Effects of HPC atrophy
Anterograde retrieval	Verbal recognition	Does not explain additional variance beyond that explained by inter-HPC rsFC reduction
	Visual Recognition	Fully mediated by PCC rsALFF reduction
	Verbal Recall
	Visual Recall
Anterograde Retention (Visual Forgetting)		Direct effect - not mediated by extra-HPC abnormalities
Remote Autobiographical Memory		Fully mediated by thalamic volume reduction

HPC: hippocampus; PCC: posterior cingulate cortex; rsALFF: resting-state amplitude of low frequency fluctuations; rsFC: resting-state functional connectivity.

Discussion

This study provides one of the largest investigations to date into the neural basis of ‘hippocampal amnesia’. We examined the brain abnormalities underlying episodic memory impairment in autoimmune LE, a clinical syndrome in which acute, focal inflammation in the HPC leads, in the long-term, to HPC atrophy and amnesia. We hypothesized that HPC damage would be accompanied by remote effects on the structure and function of the extended HPC system (Aggleton et al., 2010), and that these abnormalities would be instrumental in explaining patients’ anterograde and retrograde amnesia.

Consistent with previous investigations in smaller cohorts (Butler et al., 2014; Finke et al., 2017; Henson et al., 2016; Irani et al., 2013; Malter et al., 2014), our patients showed selective deficits in episodic memory, in the face of normal visuomotor, language, executive function, general and personal semantic memory performance. Episodic memory impairment was evident across a broad range of tests that have been extensively used in studies of MTL amnesia (Bayley et al., 2003; Vann et al., 2009). In terms of anterograde memory, patients showed impaired visual and verbal recall and recognition memory, and pronounced forgetting of visual material. Examining retrograde amnesia, we found striking loss of remote autobiographical memories from childhood and early adulthood.

The focal HPC high T2 signal on clinical MRI from the acute disease phase was followed by pronounced HPC atrophy and less pronounced right entorhinal cortical volume reduction within the MTL. However, patients also showed volume reduction in specific midline and lateral thalamic regions, along with reduced rsALFF in the PCC and precuneus, and reduced rsFC between the right HPC and the left HPC, the medial prefrontal and posteromedial cortices - regions known to interact closely with the HPC in the healthy brain.

These wider network abnormalities were associated with memory performance over and above HPC atrophy, and also fully mediated the relationships between HPC atrophy and memory performance. The only direct effect of HPC atrophy we observed was on forgetting, and no other brain abnormalities showed such an effect. Our results highlight the need to take into account remote changes in brain structure and function associated with HPC damage (Aggleton, 2014), since these may explain specific aspects of anterograde and retrograde amnesia and help identify others that may be a direct function of HPC atrophy per se.

Abnormalities in the extended HPC system following HPC atrophy

We found bilaterally marked HPC atrophy and less significant right entorhinal cortical volume reduction using the gold standard of manual volumetry for MTL structures (Grimm et al., 2015). Automated segmentation of other subcortical structures disclosed left thalamic volume reduction.

However, our uniquely large cohort also enabled us to examine subtle structural abnormalities across the whole brain in a voxel-wise fashion. Using this method, volume reduction was observed in anterior-dorsomedial and dorsolateral thalamic regions, the nuclei within which are known to interact with the HPC and cingulate cortices (Aggleton et al., 2010). Our cross-sectional design cannot conclusively show that these extra-HPC abnormalities occurred as a consequence of HPC damage, an issue that needs to be addressed by future longitudinal studies. Nevertheless, acute inflammatory changes appeared confined to the HPC in the vast majority of our patients, while no such changes were found in the thalamus. Moreover, clinical T2-weighted MRIs in other autoimmune LE cohorts (Finke et al., 2017; Kotsenas et al., 2014; Malter et al., 2014), and the results of post-mortem studies have also demonstrated focal, acute HPC pathology in autoimmune LE patients (Khan et al., 2009; Park et al., 2007) and animal models (Tröscher et al., 2017). Our interpretation of these data is, therefore, that extra-HPC structures within the extended HPC system (thalamus, PCC) are affected as a secondary consequence of HPC damage, rather than due to the primary pathology. We also note that extra-HPC damage is equally if not more likely in other conditions associated with HPC amnesia, such as cases of ischemia/anoxia (Huang and Castillo, 2008). It should, however, be acknowledged that a single case study of amnesia due to ischemia/anoxia which came to post mortem demonstrated focal neuronal loss in the CA1 region of the HPC on histopathology (Zola-Morgan et al., 1986). Direct comparisons between autoimmune LE and amnesia due to different etiologies would be useful in future research.

Patients also showed functional abnormalities in the posteromedial cortex, and the relationship between HPC atrophy and PCC rsALFF was fully mediated by the correlative volume reduction in the thalamic regions. This is consistent with the functional abnormalities previously observed in posteromedial cortical regions in both HPC and diencephalic amnesias in humans (Aggleton, 2014; Reed et al., 1999). Patients also showed reduced inter-HPC rsFC. In healthy individuals, the spontaneous activity of the HPC is coupled with that of the contralateral HPC at rest and these regions form part of the default-mode network. This is also the case for the medial prefrontal and posteromedial cortical regions which showed reduced rsFC with the HPC (Buckner et al., 2008; Greicius et al., 2004). The decrease in inter-HPC rsFC across patients as a function of delay since symptom onset is consistent with the idea that these network-wide abnormalities follow focal HPC damage. In future, this needs replication in longitudinal studies.

Anterograde memory: Retrieval

Patients showed impairment across all tests of visual and verbal recall and recognition memory. The only unimpaired form of memory was face recognition. This would be predicted by some material-specific accounts of recognition memory (Bird and Burgess, 2008) and is consistent with the view that recognition of unfamiliar faces is exceptional in several regards – according to some approaches, faces may be holistically processed and difficult to label verbally, and adequate performance levels may be attained in the absence of capacities to associate faces with a study list (Smith et al., 2014). Ours is the largest study to date to confirm preserved face recognition memory in HPC amnesia.

At uncorrected levels, HPC volumes correlated with both recall and recognition memory scores. Crucially, however, the reduction in PCC rsALFF correlated with verbal and visual recall and visual recognition over and above HPC atrophy and, moreover, fully mediated the effects of HPC atrophy on these aspects of episodic memory. This finding demonstrates that the HPC role in recall and recognition cannot be examined in isolation from remote effects of HPC damage within the extended HPC system. It furthermore dovetails with the causal role that has been attributed to the posteromedial cortex in episodic memory for both visual (Bonnì et al., 2015) and verbal (Koch et al., 2018) material, and aligns with evidence from neurodegenerative diseases that it is posteromedial cortex rather than HPC function that predicts episodic memory impairment (La Joie et al., 2014). Patients’ impaired verbal recognition was associated with reduced inter-HPC rsFC. The reasons why the correlates of verbal recognition were different from those of other forms of memory are unclear, but the relationship with inter-HPC rsFC is consistent with that found between inter-HPC connectivity and episodic memory in healthy (Wang et al., 2010) and patient populations (e.g. traumatic axonal injury) (Marquez de la Plata et al., 2011).

It should be noted that our neuropsychological assessment did not explicitly attempt to distinguish between the recollection and familiarity processes that may underlie differences in recall and recognition memory. In order to identify the neural mechanisms supporting these processes, finer-grained behavioral paradigms are required. Furthermore, our study did not include HPC subfield volumetry, which may have disclosed stronger correlations between HPC subfields and memory scores. The volumes and hemodynamic activity of HPC subfields have been associated with various forms of memory (Bonnici et al., 2013; Miller et al., 2017; Palombo et al., 2018). While imaging at higher field strength is required to investigate HPC subfields, our study highlights the fact that the remote effects of HPC damage on wider brain networks are a key factor in HPC amnesia.

It is important to acknowledge an alternative interpretation of the relationship that we observed between resting-state functional abnormalities and memory measures. McCormick and colleagues recently showed that HPC amnesic patients differ in the form and content of their ‘mind-wandering’ compared with healthy adults (McCormick et al., 2018). It is possible that ‘resting-state’ functional abnormalities in our patients actually reflect (and are perhaps therefore mediated by) differences between healthy controls and patients with respect to the extent of mind wandering in the scanner, rather than differences in neurovascular functioning in these regions per se. Anterior and posterior midline structures are strongly implicated in mind-wandering. Nevertheless, rsfMRI measures (predominantly rsFC) have repeatedly provided reliable correlates of memory impairment in HPC amnesia [e.g. (Heine et al., 2018; Henson et al., 2016)]. Moreover, this interpretation is not inconsistent with the basic premise of our argument, namely, that the effects of HPC damage on memory are mediated by other processes that are compromised following HPC damage. Our study was not designed to disambiguate the level of disruption that is mediating the effects of HPC atrophy. In other words, the disruption could be at the cognitive level, or at the neurovascular level, or at both levels. Further work is needed to disambiguate those two interpretations.

Anterograde memory: Forgetting

The only aspect of amnesia upon which HPC atrophy showed a direct effect was that of visual forgetting (of abstract shapes). Patients showed only a marginal increase in verbal forgetting. In particular, it was the atrophy in the right HPC that was associated with visual forgetting. This is consistent with evidence for HPC lateralization of maintenance processes for verbal (left HPC) (Frisk and Milner, 1990) and visual material (right HPC) (Smith and Milner, 1989). Indeed, rapid forgetting of visual information in right HPC damage was noted quite early (Jones-Gotman, 1986), as was, in general, the nature of forgetting in HPC amnesia (Huppert and Piercy, 1979). This has recently been re-emphasized (Sadeh et al., 2014), with functional neuroimaging studies in healthy young adults attributing a central role to the HPC in constraining the forgetting that occurs by new learning (Kuhl et al., 2010).

Our results go beyond these studies and demonstrate an exclusive relationship between HPC atrophy and rapid forgetting even within the context of abnormalities in the extended HPC system. This relationship is consistent with computational models of HPC function that outline pattern completion and pattern separation (O'Reilly and McClelland, 1994) as two core mechanisms that protect against forgetting. Pattern completion enables the reinstatement of previously encoded memories from incomplete input, whereas pattern separation involves the orthogonal coding of memories for overlapping events, which minimizes forgetting by generating non-interfering representations (Kuhl et al., 2010).

These findings suggest that the HPC role in amnesia may be much less direct than has previously been held, at least with respect to performance on widely used, standardized neuropsychological tests of memory. In contrast with the majority of such tests, the measurements of forgetting such as that used here partially control for factors that may confound simple recall scores, such as variability in attention, initial learning or retrieval strategy, factors that may depend on regions outside the HPC. Instead, measurements of forgetting may provide the most sensitive and specific way to detect HPC damage. This proposal does not preclude the possibility that the HPC mechanisms underlying the retention of newly formed memories are process- or material-specific. As our forgetting rates were derived only from a recall-based test (Baddeley et al., 1994), we are unable to comment on the relationship of HPC atrophy with forgetting in recognition memory, a relationship which has recently been questioned (Sadeh et al., 2014). Likewise, material-specificity in rapid forgetting should be examined with finer-grained behavioral tasks, involving different types of stimuli (e.g. scenes vs. faces). Nevertheless, the direct relationship of (right) HPC atrophy with (visual) forgetting highlights the utility of tests of accelerated forgetting in detecting HPC pathology in other disorders. Recent studies have emphasized a possible role for such tests in the early detection of Alzheimer's disease pathology (Weston et al., 2018; Zimmermann and Butler, 2018).

Remote autobiographical memory

Patients showed spared remote personal semantic memory, in the face of impaired remote autobiographical memories. To date, research in autoimmune LE patients has focused on anterograde memory (Butler et al., 2014; Finke et al., 2017; Henson et al., 2016; Malter et al., 2014; Miller et al., 2017), with very little evidence on retrograde amnesia (Chan et al., 2007). Our study is also one of the very few to examine the relationship of HPC damage with both anterograde and retrograde amnesia.

It is possible that the test we used to assess remote memories (AMI) is insufficiently sensitive to truly ‘episodic’ aspects of autobiographical memory that have been held to be HPC-dependent (Moscovitch et al., 2016; Nadel and Moscovitch, 1997). Other methods, involving parametric text-based procedures, may offer greater sensitivity [e.g. the Autobiographical Interview (Levine et al., 2002)]. Nevertheless, studies on other amnesic cohorts have found the AMI and the Autobiographical Interview to offer comparable results (Rensen et al., 2017).

Our findings would at first sight appear to support a role for the HPC in the recollection of remote autobiographical memories. However, the volume of the left thalamus correlated with remote autobiographical memory scores over and above HPC volumes across patients. Moreover, the effects of HPC atrophy were fully mediated by the volume reduction in the thalamus. Furthermore, no selective relationship was found between remote autobiographical memory scores and anterior or posterior HPC volumes. These findings are difficult to reconcile with the multiple trace (Moscovitch et al., 2005) or trace-transformation accounts of retrograde amnesia (Sekeres et al., 2018), according to which remote autobiographical memory deficits should be a function of the extent of HPC damage, either in toto, or in specific HPC portions.

The absence of direct effects of HPC atrophy on remote memory, in combination with the selective presence of such effects on forgetting rates, is instead consistent with the idea that HPC circuitry acts as a buffer for the temporary maintenance of episodic information, with non-HPC structures storing more permanent memory traces. Our findings may thus offer some support for the standard consolidation model of remote memory. Other work supporting this model has primarily emphasized the role of neocortical damage in retrograde amnesia (Bayley et al., 2003; Squire et al., 2015). We here highlight the need also to consider the role of the thalamus. Focal thalamic damage has been often associated with retrograde amnesia in single-case or small case-series studies (Carlesimo et al., 2011; Stuss et al., 1988), and mediodorsal thalamic nuclei may coordinate the recollection of cortically stored memories (Miller et al., 2001) via thalamic-prefrontal cortical projections (Kopelman, 2015). Our results show that secondary alterations in thalamic integrity may explain the large variability in retrograde memory loss reported after HPC damage (Bartsch et al., 2011; Bayley et al., 2003; St-Laurent et al., 2011).

Finally, our finding that thalamic volume was reduced in our patient group is consistent with the literature on developmental amnesia due to early hypoxic-ischaemic encephalopathy, where HPC damage has been noted along with atrophy in the thalamus and the mammillary bodies [e.g. (Dzieciol et al., 2017)]. A question for future research is therefore whether adult-onset HPC damage is accompanied by atrophy in the mammillary bodies, or whether thalamic atrophy may occur in the absence of changes to the mammillary bodies, given the evidence for direct forniceal projections from the HPC/subiculum to the anterior thalamus in human and non-human primates (Bubb et al., 2017).

Conclusion

Our study, probably the largest in human HPC amnesia, shows that abnormalities in the integrity of and connectivity within the extended HPC system may occur after focal HPC damage, and that these play a central role in explaining the variability in the anterograde and retrograde amnesia of patients with HPC atrophy. We found that the neuropsychological measure most sensitive and specific to HPC atrophy was the forgetting of newly learned information. Understanding the impact of network-wide changes following HPC damage will be key to resolving long-standing controversies in the literature on human amnesia.

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Outline of Results Section.

Neuropsychological profile of autoimmune LE patients (post-acute phase) and healthy controls

Clinical details of autoimmune LE patients (acute phase).

Volumetry of MTL and subcortical structures in autoimmune LE patients.

Reduction in GM volume, rsALFF and rsFC in autoimmune LE patients.

GM volume reduction in autoimmune LE patients (whole-brain VBM).

Verbal Recognition Memory: Structural/Functional correlates.

Figure 3—source data 1

Visual Recognition Memory: Structural/Functional correlates.

Figure 4—source data 1

Verbal Recall Memory: Structural/Functional correlates.

Figure 5—source data 1

Visual Recall Memory: Structural/Functional correlates.

Figure 6—source data 1

Forgetting: Structural/Functional correlates.

Figure 7—source data 1

Remote Autobiographical Memory: Structural/Functional correlates.

Figure 8—source data 1

Summary of relationships between memory impairment and structural/functional abnormalities across patients.

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Georgios PD Argyropoulos

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Clare Loane

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Adriana Roca-Fernandez

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Carmen Lage-Martinez

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Oana Gurau

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Sarosh R Irani

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Christopher R Butler

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