Generality and opponency of rostromedial tegmental (RMTg) roles in valence processing

  1. Hao Li
  2. Dominika Pullmann
  3. Jennifer Y Cho
  4. Maya Eid
  5. Thomas C Jhou  Is a corresponding author
  1. Medical University of South Carolina, United States
6 figures


Behavioral training procedure.

(A) Schematic of training paradigm. (B) Training performance. Response index is the percentage of trials in which animals made nose pokes within 2 s of onset of reward or neutral cues.
Figure 2 with 1 supplement
RMTg neurons are activated by diverse phasic aversive stimuli.

(A) Schematic of recording paradigm. (B) Raster plots of a representative RMTg neuron response to reward cues and footshocks. (C) RMTg neurons on average showed inhibition to reward-predictive cues during a 200–400 ms post-stimulus window, and small excitations to neutral tones and large excitations to footshock, siren, and bright light during a 0–100 ms post-stimulus windows. (D) Percentage of RMTg neurons that showed inhibition, excitation or no response to stimuli (200–400 ms window for reward cues, and 0–100 ms window for aversive or neutral stimuli). (E) Scatterplot of individual neurons’ responses to reward cues and footshocks. Many reward-cue inhibited neurons were also excited by footshocks, consistent with a valence-encoding pattern. Blue solid dots: neurons significantly responding to both reward cues and footshocks. Gray shaded box: neurons inhibited by reward cues and excited by footshocks, consistent with hypothesized valence-encoding. (F, G) RMTg neurons activated by footshocks tended to also be activated by siren and bright light, in proportion to the magnitude of response to footshock. Solid dots: neurons significantly responding to siren and bright light. * indicates p < 0.05, ** p < 0.01, *** < 0.0001.
Figure 2—figure supplement 1
Histology of RMTg recordings.

(A) Photo of an electrode track in the RMTg region stained with FOXP1 (black dots). Red dashed line delineates RMTg as the region of dense FOXP1 expression. Black line: electrode. (B) Recording sites in the RMTg region verified by FOXP1 staining.
RMTg neurons exhibit biphasic responses to sustained aversive stimuli consistent with opponent process theory.

(A, B) RMTg neurons showed activation to a low dose of LiCl for 10 min post injection, and (E, F) to restraint stress during the first 3 min of the 6 min restraint. Both aversive stimuli also produced a rebound inhibition of firing below baseline during a later time window (20–30 min window for LiCl, and 9–12 min window for restraint stress). (C, D) Individual neuron responses during initial phases of LiCl and restraint stress correlated with their responses to footshock, while responses during rebound phases correlated with responses to reward cue (G, H). (I, J) Cocaine infusion produced an opposing pattern in RMTg neurons, with inhibition during the first 10 min followed by a rebound excitation 15–25 min post infusion. (K, L) Individual responses during rebound (aversive) phase of cocaine were correlated with their responses to footshock, while responses during initial (rewarding) phase were correlated with their responses to reward cue. (M) Schematic of conditioned place preference regimen. (N) Low dose of LiCl (10 mg/kg) i.p. injection induced place aversion during 0–15 min and place preference during 15–30 min post-stimulus. (O) Cocaine (0.75 mg/kg) i.v. infusion induced place preference during 0–15 min and place aversion 15–30 min post-stimulus. (P) Saline injection did not produce place preference during either time window. Solid dots in B, F, and J: neurons significantly responding during both initial and rebound phases.
Figure 4 with 1 supplement
VTA-projecting RMTg neurons preferentially show valence-encoding patterns.

(A) Cav2-cre injected into the VTA or DRN was retrogradely transported to the RMTg in mice, driving gCaMP6f expression in subsets of RMTg neurons projecting to the VTA or DRN respectively. (B) Representative photograph of the RMTg region in which gCaMP6f in RMTg (green label) is co-expressed with FOXP1 (red), a transcription factor locally specific to RMTg neurons. (C) A representative photo of gCaMP6f positive neurons in vivo (upper panel) and denoised Ca2+ traces extracted from the marked neurons (lower panel). (D) VTA-projecting RMTg neurons showed an average inhibition by reward cues, and excitation by footshocks. (E) DRN-projecting neurons showed no average response to reward cues, but were excited by footshocks. (F) Among VTA-projecting RMTg neurons, neurons showing stronger excitations to shock tended to also show stronger inhibitions to the reward cue, while individual DRN neurons did not show this correlation (G). Colored solid dots: neurons significantly responding to both reward cues and footshocks. (H) VTA-projecting neurons were much more likely to be inhibited by the reward cue than DRN-projecting neurons, and much less likely to be activated, while VTA- and DRN-projecting neurons were both predominantly activated by shock.
Figure 4—figure supplement 1
Histology of endoscopic calcium imaging and individual responses of VTA-projecting RMTg neurons.

(A) Photo of GRIN lens track in the RMTg. Green cells are gCaMP6f-positive cells. (B) Lens placements (Black: VTA-projecting group; Gray: DRN-projecting group). (C, D) Calcium signals (dF/F) from individual VTA-projecting RMTg neurons in response to reward cues and footshocks. Black bar: duration of the cue; Blue circle: sucrose delivery; Red star: onset of footshocks.
Figure 5 with 2 supplements
Excitotoxic RMTg lesions abolished VTA inhibitions by aversive stimuli.

(A, B) Heatmap showing all VTA neuron responses to reward cues in sham group. pDA neurons in the VTA were classified by their phasic activation to reward cues (0–200 ms post-stimulus window), while pGABA neurons were classified by the presence of sustained activations (200–2000 ms post-stimulus window). (C) Raster plots of immediate inhibition and delayed inhibition response types observed in pDA neuron after aversive stimuli. (D) Comparisons of RMTg (blue trace) and pDA (red trace) neuron responses to affective stimuli. All three aversive stimuli elicited initial excitations in both RMTg and pDA neurons, after which RMTg neurons remained excited while pDA neurons showed inhibition during 100–500 ms window post-stimulus (brown-shaded boxes). pDA neurons were activated by reward cues, and at faster latencies than RMTg inhibition to the same cue, making it unlikely that pDA activations to the reward cue would be driven by the RMTg. (E) NeuN staining showed that lesions were mostly restricted to the RMTg area and did not extend to surrounding structures such as the pedunculopontine nucleus (PPTg) and dorsal raphe nucleus (DRN). Scalebars: 1 mm and 100 μm for left and right panels. (F) RMTg lesion (dashed trace) eliminated aversion-induced inhibition in pDA neurons. (G) Bar graphs again showing loss of aversion-induced inhibition during 100–500 ms in pDA neurons after RMTg lesions. (H) Loss of aversion-induced inhibition in all recorded VTA neurons.
Figure 5—figure supplement 1
Histology of VTA recordings.

(A) Photo of an electrode track in the VTA region stained with TH. Red dashed line: the shape of the VTA. Black line: electrode. (B) Recording sites in the VTA region verified by TH staining in intact rats (blue dots) or lesioned rats (red dots).
Figure 5—figure supplement 2
Supplementary VTA recordings. 

In unlesioned animals, pDA neurons responded to aversive stimuli with either pure inhibitionn (A), inhibition after a brief excitation (B), or pure excitation (C). The percentages of neurons responding to different aversive stimuli in each category are listed in each panel. (D) RMTg lesions did not alter percentages of pDA neurons with excitatory/inhibitory responses to reward/neutral tones. (E) RMTg lesions did not alter pDA responses to reward cue. (F) RMTg lesions did not change basal firing , but increased the percentage of pDA spikes in bursts. (G) RMTg lesions slightly increased the proportion of pDA neurons excited by aversive stimuli during 100–500 ms post-stimulus. (H) The percentages of pDA neurons with initial excitations to aversive stimuli were unaffected after RMTg lesion, while the magnitudes of these initial excitations were significantly increased.
Figure 6 with 1 supplement
RMTg lesions disrupt conditioned place aversion to a wide range of stimuli.

(A) Photographs of immunostaining of NeuN in RMTg region with and without excitotoxic lesions. Scalebar: 100 µm. RMTg-lesions reduced place aversion scores as measured by relative time spent in stimulus-paired versus unpaired chambers (B), and abolished place aversion as measured by relative entries into paired versus unpaired chambers (C). (D) RMTg lesions did not alter total entries into chambers. (E) RMTg-lesioned rats lacked the training-induced reduction in locomotion seen in shams (F, G).
Figure 6—figure supplement 1
Excitotoxic lesion placements in the RMTg.

RMTg (black circles) lesion sizes (yellow) in conditioned place aversion experiments.

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  1. Hao Li
  2. Dominika Pullmann
  3. Jennifer Y Cho
  4. Maya Eid
  5. Thomas C Jhou
Generality and opponency of rostromedial tegmental (RMTg) roles in valence processing
eLife 8:e41542.