Subcortico-amygdala pathway processes innate and learned threats
Curation statements for this article:-
Curated by eLife
eLife assessment
This study presents valuable insights into the circuits that are common for innate and learned threat. The evidence supporting the author's conclusions is solid, but the specificity of the circuit targeting methods requires further histological assessment and clarification. Deeper interpretation of novel mechanistic insights that are gained would benefit the study.
This article has been Reviewed by the following groups
Listed in
- Evaluated articles (eLife)
- Neuroscience (eLife)
Abstract
Behavioral flexibility and timely reactions to salient stimuli are essential for survival. The subcortical thalamic-basolateral amygdala (BLA) pathway serves as a shortcut for salient stimuli ensuring rapid processing. Here, we show that BLA neuronal and thalamic axonal activity in mice mirror the defensive behavior evoked by an innate visual threat as well as an auditory learned threat. Importantly, perturbing this pathway compromises defensive responses to both forms of threats, in that animals fail to switch from exploratory to defensive behavior. Despite the shared pathway between the two forms of threat processing, we observed noticeable differences. Blocking β-adrenergic receptors impairs the defensive response to the innate but not the learned threats. This reduced defensive response, surprisingly, is reflected in the suppression of the activity exclusively in the BLA as the thalamic input response remains intact. Our side-by-side examination highlights the similarities and differences between innate and learned threat-processing, thus providing new fundamental insights.
Article activity feed
-
-
Author Response
Reviewer #2 (Public Review):
"... the fact that MGN-BLA circuit disruptions were done during the conditioning phase of associative threat learning, and not during the recall phase only, complicates the side-by-side comparison: it could be argued that in this case what is disturbed is the processing of the unconditioned innately aversive stimulus in the task, the foot shock, instead of the learnt threat of the sound".
In our previous email to the editors, we mentioned work by Barsy et al., showing that indeed the inhibition of this input during the recall phase reduces freezing response (Please see Fig. 8 in Barsy et al). In the new revision, we refer to this experiment.
Specific comments (weaknesses):
e) There are not enough analysis and method descriptions to demonstrate the specificity of the targeting approach
Author Response
Reviewer #2 (Public Review):
"... the fact that MGN-BLA circuit disruptions were done during the conditioning phase of associative threat learning, and not during the recall phase only, complicates the side-by-side comparison: it could be argued that in this case what is disturbed is the processing of the unconditioned innately aversive stimulus in the task, the foot shock, instead of the learnt threat of the sound".
In our previous email to the editors, we mentioned work by Barsy et al., showing that indeed the inhibition of this input during the recall phase reduces freezing response (Please see Fig. 8 in Barsy et al). In the new revision, we refer to this experiment.
Specific comments (weaknesses):
e) There are not enough analysis and method descriptions to demonstrate the specificity of the targeting approach
We have included these data as supplementary figures (S2A and B, S5B, S7, S9A and S10K) and added a more detailed methodology in the method section.
f) …the authors administer blockers of beta-adrenergic receptors systemically. This reveals differences between MGN-BLA projecting neurons, BLA neurons, and innate and learnt threat, but the mechanistic implications are not clear and should be discussed.
In the revised manuscript, we extensively discuss these points: (This indicates that the looming stimulus conveyed through the thalamic input…may contribute to the variability in the effect of the drug in freezing response); (...in mice injected with propranolol, the defensive responses…The differences in species or strains used, or experimental parameters may contribute to the variability in the effect of the drug in freezing response.)
-
eLife assessment
This study presents valuable insights into the circuits that are common for innate and learned threat. The evidence supporting the author's conclusions is solid, but the specificity of the circuit targeting methods requires further histological assessment and clarification. Deeper interpretation of novel mechanistic insights that are gained would benefit the study.
-
Reviewer #1 (Public Review):
The authors expand upon prior findings and show that basolateral amygdala (BLA) activity is necessary for defensive responses elicited by both innate and learned threats. The authors also show that a projection from the auditory thalamus (MGM thalamic nucleus) mediates these effects.
Learned threats were modelled with auditory fear conditioning. The authors finding showing that the MGM-BLA pathway is required for auditory fear learning is largely a replication of prior results.
The novelty in this paper is that the authors show that the auditory MGM-BLA pathway is involved in defense evoked by a visual looming stimulus.
Overall, this is a reasonably designed study. The main weakness is that the loss of function manipulations use either caspase-induced lesions or contralateral chemogenetic disconnection …
Reviewer #1 (Public Review):
The authors expand upon prior findings and show that basolateral amygdala (BLA) activity is necessary for defensive responses elicited by both innate and learned threats. The authors also show that a projection from the auditory thalamus (MGM thalamic nucleus) mediates these effects.
Learned threats were modelled with auditory fear conditioning. The authors finding showing that the MGM-BLA pathway is required for auditory fear learning is largely a replication of prior results.
The novelty in this paper is that the authors show that the auditory MGM-BLA pathway is involved in defense evoked by a visual looming stimulus.
Overall, this is a reasonably designed study. The main weakness is that the loss of function manipulations use either caspase-induced lesions or contralateral chemogenetic disconnection studies, which lack temporal resolution.
-
Reviewer #2 (Public Review):
Khalil et al. aimed to gain insights into similarities and differences between circuits processing innate and learned threats. For this, they investigated a circuit that is well established to have a critical role in auditory associative threat learning, the projection from the medial geniculate nucleus (MGN) to the basolateral amygdala (BLA), and carried out a side-by-side comparison of its role in conditioned and innate threat.
Although the MGN is part of the main auditory stream, the neurons that project to BLA are multimodal. Khalil et al. took advantage of this to use visual looming stimuli to evoke innate threat. The authors showed that the MGN-BLA pathway processes both innate freezing responses to looming black circles and threat-conditioned freezing responses to tones. The disruption of the pathway …
Reviewer #2 (Public Review):
Khalil et al. aimed to gain insights into similarities and differences between circuits processing innate and learned threats. For this, they investigated a circuit that is well established to have a critical role in auditory associative threat learning, the projection from the medial geniculate nucleus (MGN) to the basolateral amygdala (BLA), and carried out a side-by-side comparison of its role in conditioned and innate threat.
Although the MGN is part of the main auditory stream, the neurons that project to BLA are multimodal. Khalil et al. took advantage of this to use visual looming stimuli to evoke innate threat. The authors showed that the MGN-BLA pathway processes both innate freezing responses to looming black circles and threat-conditioned freezing responses to tones. The disruption of the pathway impairs freezing in both cases, and the pathway is activated mostly in the presence of freezing. This suggests that the MGN-BLA processes threat independently of the sensory modality and of whether the threat is learnt or not. This further suggests that these different forms of threat may share similar mechanisms.
Nonetheless, the fact that MGN-BLA circuit disruptions were done during the conditioning phase of associative threat learning, and not during the recall phase only, complicates the side-by-side comparison: it could be argued that in this case what is disturbed is the processing of the unconditioned innately aversive stimulus in the task, the foot shock, instead of the learnt threat of the sound. Still, this would go in hand with one of the main conclusions of the study, which is that the MGN-BLA processes innate threats.
There are alternative interpretations of the results though, which are beyond the scope of the study: the circuit might be relevant for processing salient stimuli beyond threatening stimuli, for instance for positive valence stimuli as well; or this circuit might be relevant for processing the freezing response to threat in particular. To target the MGN-BLA circuit, the authors employ viral-vector mediated expression of proteins in mice. This way they delete, inhibit, or image either the activity of the neurons (or the axons) that project from MGN to BLA, or the BLA neurons themselves. They combine this with fiber-photometry and behavioural quantifications. Targeting these small and deep nuclei in the mouse brain bilaterally is challenging, which increases the value of the presented data. Conversely, it is important that the authors support more explicitly the specificity of their targeting methods and quantifications throughout the manuscript.
Overall, the main conclusions of this paper are mostly supported by data, but important methodological aspects need to be clarified, data analysis extended and the interpretation of results discussed further. The question of whether innate and learnt responses to stimuli share common mechanisms is timely. This study places the MGN-BLA pathway as a suitable model circuit to investigate this and paves the way for future work to dig into the implicated mechanisms.
Specific comments (strengths):
a) The authors use two methods to interrupt the MGN-BLA pathway, a reversible one (chemogenetics) and an irreversible one (neuronal deletion via caspase 3 expression), obtaining consistent results that strengthen the evidence supporting their conclusions.
b) The authors demonstrate the efficacy of their MGN-BLA pathway interruption methods with in vivo recordings.
c) The approach of addressing the same behavioural output (freezing) in the two conditions (innate and learnt threat) helps the interpretability of results.Specific comments (weaknesses):
e) There are not enough analysis and method descriptions to demonstrate the specificity of the targeting approach, which is in some cases neither reflected in the pictures of the main figures. These include quantifications of the extension of expression/deletions in the brain and placement of viral-vector injections. In particular, these should show that i) protein expression does not extend beyond the BLA or MGN; ii) the MGN cells projecting to the striatum (right above the BLA) are not implicated, iii) that neurons in the visual thalamus are not affected by the manipulations. These are critical points that need to be addressed.
f) There is a lack of digging into the mechanisms that could be enhanced with further analysis and discussion. For example, to start addressing this question, the authors administer blockers of beta-adrenergic receptors systemically. This reveals differences between MGN-BLA projecting neurons, BLA neurons, and innate and learnt threat, but the mechanistic implications are not clear and should be discussed. Also, the interpretation of the pathway's role in behaviour and its relation to neuronal activity could be deepened with further analysis. -
Reviewer #3 (Public Review):
Khalil et al. investigated the role of medial geniculate nuclei -> basolateral amygdala pathway in the processing of innate and learned threats. Using looming stimuli and cued fear conditioning the authors show that both the BLA and MGN projections to the BLA respond to learned and innately threatening stimuli and that their activation is necessary to generate adequate fear responses. Lastly, Khalil et al. highlight a possible role of adrenergic signaling in modulating threat-induced BLA (but not MGN) activity. The manuscript is well conceived, the statistical analysis is solid, and the methodology is appropriate. The strength of this paper is that the hypothesis is tested using multiple experimental strategies that all nicely converge to demonstrate the involvement of the MGN-BLA pathway in threat …
Reviewer #3 (Public Review):
Khalil et al. investigated the role of medial geniculate nuclei -> basolateral amygdala pathway in the processing of innate and learned threats. Using looming stimuli and cued fear conditioning the authors show that both the BLA and MGN projections to the BLA respond to learned and innately threatening stimuli and that their activation is necessary to generate adequate fear responses. Lastly, Khalil et al. highlight a possible role of adrenergic signaling in modulating threat-induced BLA (but not MGN) activity. The manuscript is well conceived, the statistical analysis is solid, and the methodology is appropriate. The strength of this paper is that the hypothesis is tested using multiple experimental strategies that all nicely converge to demonstrate the involvement of the MGN-BLA pathway in threat processing. However, a more detailed analysis of fiber photometry data in relation to the presented stimuli and to behavioral responses would help to clarify whether this MGN-BLA pathway is involved in processing sensory stimuli per se or directly generates behavioral responses.
-
