Multisensory integration enhances audiovisual responses in the Mauthner cell

Reviewer #1 (Public Review):

Summary:

Otero-Coronel et al. address an important question for neuroscience - how does a premotor neuron capable of directly controlling behavior integrate multiple sources of sensory inputs to inform action selection? For this, they focused on the teleost Mauthner cell, long known to be at the core of a fast escape circuit. What is particularly interesting in this work is the naturalistic approach they took. Classically, the M-cell was characterized, both behaviorally and physiologically, using an unimodal sensory space. Here the authors make the effort (substantial!) to study the physiology of the M-cell taking into account both the visual and auditory inputs. They performed well-informed electrophysiological approaches to decipher how the M-cell integrates the information of two sensory modalities depending on the strength and temporal relation between them.

Strengths:

The empirical results are convincing and well-supported. The manuscript is well-written and organized. The experimental approaches and the selection of stimulus parameters are clear and informed by the bibliography. The major finding is that multisensory integration increases the certainty of environmental information in an inherently noisy environment.

Weaknesses:

Even though the manuscript and figures are well organised, I found myself struggling to understand key points of the figures.

For example, in Figure 1 it is not clear what are actually the Tonic and Phasic components. The figure will benefit from more details on this matter. Then, in Figure 4 the label for the traces in panel A is needed since I was not able to pick up that they were coming from different sensory pathways.

In line 338 it should be optic tectum and not "optical tectum".

Reviewer #2 (Public Review):

Summary:

In this manuscript, Otero-Coronel and colleagues use a combination of acoustic stimuli and electrical stimulation of the tectum to study MSI in the M-cells of adult goldfish. They first perform a necessary piece of groundwork in calibrating tectal stimulation for maximal M-cell MSI, and then characterize this MSI with slightly varying tectal and acoustic inputs. Next, they quantify the magnitude and timing of FFI that each type of input has on the M-cell, finding that both the tectum and the auditory system drive FFI, but that FFI decays more slowly for auditory signals. These are novel results that would be of interest to a broader sensory neuroscience community. By then providing pairs of stimuli separated by 50ms, they assess the ability of the first stimulus to suppress responses to the second, finding that acoustic stimuli strongly suppress subsequent acoustic responses in the M-cell, that they weakly suppress subsequent tectal stimulation, and that tectal stimulation does not appreciably inhibit subsequent stimuli of either type. Finally, they show that M-cell physiology mirrors previously reported behavioural data in which stronger stimuli underwent less integration.

The manuscript is generally well-written and clear. The discussion of results is appropriately broad and open-ended. It's a good document. Our major concerns regarding the study's validity are captured in the individual comments below. In terms of impact, the most compelling new observation is the quantification of the FFI from the two sources and the logical extension of these FFI dynamics to M-cell physiology during MSI. It is also nice, but unsurprising, to see that the relationship between stimulus strength and MSI is similar for M-cell physiology to what has previously been shown for behavior. While we find the results interesting, we think that they will be of greatest interest to those specifically interested in M-cell physiology and function.

Strengths:

The methods applied are challenging and appropriate and appear to be well executed. Open questions about the physiological underpinnings of M-cell function are addressed using sound experimental design and methodology, and convincing results are provided that advance our understanding of how two streams of sensory information can interact to control behavior.

Weaknesses:

Our concerns about the manuscript are captured in the following specific comments, which we hope will provide a useful perspective for readers and actionable suggestions for the authors.

Comment 1 (Minor):

Line 124. Direct stimulation of the tectum to drive M-cell-projecting tectal neurons not only bypasses the retina, it also bypasses intra-tectal processing and inputs to the tectum from other sources (notably the thalamus). This is not an issue with the interpretation of the results, but this description gives the (false) impression that bypassing the retina is sufficient to prevent adaptation. Adding a sentence or two to accurately reflect the complexity of the upstream circuitry (beyond the retina) would be welcome.

Comment 2 (Major):

The premise is that stimulation of the tectum is a proxy for a visual stimulus, but the tectum also carries the auditory, lateral line, and vestibular information. This seems like a confound in the interpretation of this preparation as a simple audio-visual paradigm. Minimally, this confound should be noted and addressed. The first heading of the Results should not refer to "visual tectal stimuli".

Comment 3 (Major):

Figure 1 and associated text.

It is unclear and not mentioned in the Methods section how phasic and tonic responses were calculated. It is clear from the example traces that there is a change in tonic responses and the accumulation of subthreshold responses. Depending on how tonic responses were calculated, perhaps the authors could overlay a low-passed filtered trace and/or show calculations based on the filtered trace at each tectal train duration.

Comment 4 (Minor):

Figure 3 and associated text.
This is a lovely experiment. Although it is not written in text, it provides logic for the next experiment in choosing a 50ms time interval. It would be great if the authors calculated the first timepoint at which the percentage of shunting inhibition is not significantly different from zero. This would provide a convincing basis for picking 50ms for the next experiment. That said, I suspect that this time point would be earlier than 50m s. This may explain and add further complexity to why the authors found mostly linear or sublinear integration, and perhaps the basis for future experiments to test different stimulus time intervals. Please move calculations to Methods.

Comment 5 (Major):

Figure 4C and lines 398-410.
These are beautiful examples of M-cell firing, but the text suggests that they occurred rarely and nowhere close to significantly above events observed from single modalities. We do not see this as a valid result to report because there is insufficient evidence that the phenomenon shown is consistent or representative of your data.

Author response:

Answers to Reviewer #1 (Public Review):

(1) Tonic and phasic components in Figure 1 are not clear.

We will reformulate Figure 1A to show how the tonic and phasic components were measured. As this point was also raised by Reviewer #2 (Comment 3), we will explicitly clarify this in the Methods section. We will modify the color scheme to improve clarity.

(2) Labeling of traces in Figure 4.

We will add labels to traces informing which sensory pathways were stimulated to produce each response.

(3) Optic tectum instead of optical tectum.

We apologize for the error. We will replace “optical tectum” with “optic tectum” as also suggested by Reviewer #2.

Answers to Reviewer #2 (Public Review):
(1) Complexity of tectum upstream circuitry (Comments 1 and 2).

Processing of visual information is certainly a major role of the tectum, but it is true that it also receives sensory inputs from other structures including sensory pathways. We will acknowledge this complexity in our revised manuscript along with suggestions for heading titles.

(2) Figure 1 and associated text.

As mentioned in the provisional answer point 1 to Reviewer #1, we will reformulate Figure 1A and clarify how tonic and phasic responses were calculated.

(3) Figure 3 and associated text.

We will perform the analysis suggested by the reviewer and move calculations to the Methods section as requested.

(4) Figure 5C and lines 398-410.

We will consider omitting Figure 5C or clearly stating its value in the context of the rest of the data and our previous behavioral experiments.