Synapses in the Zone!

Dr. Kristen Harris and her lab have made great strides in the field of structural basis of synaptic plasticity using physiology and electron microscopy image reconstruction. In this study this lab began to explore the structure of nascent and active synapses and the mechanism behind the synaptic plasticity of recruiting these nascent synapses. It has been shown that nascent synapses lay on the outskirts of active synapses and serve as highly dynamic regions for AMPA receptor activity. Even with this receptor traffic though, these nascent synapses lack neurotransmitter vesicles and therefore stay dormant. What has not been studied is if Dense Core Vesicles (DCVs) are trafficked to these nascent synapses under Long term potentiation (LTP) conditions. DCVs are know to transport presynaptic scaffolding and many proteins necessary for vesicle docking as well as cell adhesion proteins.

In order to explore this further, the Harris lab stimulated with a Theta-burst stimulation and recorded from rat hippocampal slices in the CA1. They fixed tissue after 5min, 30min, and 2hrs. They used 3D electron microscopy (EM), EM tomography, and image reconstruction to study the structural changes in these slices. The first thing that they did was to characterize active and nascent synapses and their vesicles through EM imaging. They observed that synapses that did have nascent zones most likely had them on the outside of the active zones. Also, they show that under LTP conditions active zones undergo a larger area expansion then those without nascent zones. This begins to show nascent zone influence on active zones in the synapse. They finish by showing that more DCVs traffic to nascent zones with LTP stimulation. This in turn promotes an increase in the minimum distance necessary between docked vesicles and nascent zones for these nascent zones to be converted into active zones, as shown in the figure. This study offers a window into the mechanisms behind synaptic plasticity and illuminates some of the structural mysteries behind synaptic efficacy.

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To learn more about Dr. Harris’s research, please attend her talk on Tuesday, May 16, 2017 at 4pm in the CNCB Marilyn G. Farquhar Seminar Room.

Kevin White is a PhD student currently rotating in Dr. Axel Nimmerjan’s lab in the Biophontonics Center at the Salk Institute.


Credit where Credit’s Due

While astrocytes are the most abundant cells in the brain, their role in cognition and disease remains very poorly studied.  What is known is that they play a vital role in regulating extracellular signaling and the development and maturation of neurons.  It follows that abnormalities in glia might contribute to certain neural developmental disorders.  One such disease is Costello syndrome which is caused by mutations in the HRAS gene. Erik Ullian et al. used mutant HRAS human induced pluripotent stem cells (iPSCs) and transgenic mice to study the lineage of astroglia in this disease.

The researchers found that iPSCs from humans with Costello syndrome behaved differently than those from healthy controls in several ways. First, they differentiated into astroglia more quickly, second, they grew larger, and finally, they produced more extracellular remodeling factors and proteoglycans.  Similarly, mice who had astrocyte selective mutations in HRAS showed increased proteoglycans in cortex.  These changes likely cause the abnormal development observed in Costello syndrome.

Screen Shot 2017-05-08 at 5.37.12 PMTo confirm that the changes in astroglia were being caused by RAS signaling the researchers treated the iPSCs with farnesyl transferase inhibitor which selectively blocks RAS.  After two weeks of treatment they found that quantities of extracellular proteoglycans decreased and that the transcription factors associated with normal development were not affected.  These results offer new insight into the etiology of a rare but debilitating developmental disorder and point to a possible pharmacological therapy.

To learn more about Dr. Ullian’s research, please attend his talk on Tuesday, May 9, 2017 at 4pm in the CNCB Marilyn G. Farquhar Seminar Room.



Aβ acts independently of tau to stabilize microtubules.

The histopathological hallmark of Alzheimer’s disease (AD) is the presence of beta amyloid (Aβ) plaques and neurofibrillary tau (NFT) tangles in the postmortem analysis of brain tissue in a human with a medical history of progressive cognitive decline. Historically, the Amyloid Cascade hypothesis posits that a deposition of Aβ brain tissue triggers a series of pathological events that ultimately gives rise to the clinical symptoms and signs associated with Alzheimer’s disease, such as cognitive impairment and dementia (Karran et al., 2011).

But how can Aβ trigger these events? Dr. Francesca Bartolini from Columbia University studies how microtubules, the molecular skeleton of the cell, undergo post-translational modifications and how these changes can alter microtubule stability. Her lab explores how Aβ can alter microtubule stability, and how this may lead to subsequent collapses in dendritic spines, a common event in AD pathology.

The overarching model of how Aβ affects MT stability considers the role of hyperphosphorylated tau protein, which is a primary component of NFTs. Previous work suggests that Aβ accumulation results in tau hyperphosphorylation, which cannot bind well to MTs. Thus, MTs destabilize, leading to downstream neuronal damage. However, the role of Aβ on MT dynamics in the absence of tau has not been explored; in their recent paper, the Bartolini lab studied this relationship between Aβ and MTs.

In order to explore MT stability, Bartolini’s group leveraged the fact that stabilized MTs are detyrosinated such that a glutamic acid residue is exposed at the C-terminus (referred to as Glu MTs). Tyrosine residues on MTs, on the other hand, are a marker for dynamic MTs. Additionally, NIH3T3 cells were used to isolate the effects on MTs to simply Aβ, as this cell line lacks tau protein entirely. In brief, the Bartolini group found that, in NIH3T3 cells, exposure to neurotoxic levels of Aβ resulted in increased Glu MTs in a RhoA/mDia 1 dependent manner. The RhoA/mDia 1 pathway that leads to stabilization of MTs is as follows: RhoA is a Rho-GTPase that activates the formin mDia1, a protein that has primarily been studied in the context of actin dynamics. It has also been shown to be involved in MT stabilization, but the role of mDia1 on MT dynamics is still largely unclear (Chesarone et al., 2010).

In the figure below, NIH3T3 cells were exposed to soluble Aβ and fixed 2 hours later; immunostaining against detyrosinated Glu MTs revealed that 0.5-1 μM Aβ exposure lead to elevated Glu MTs compared to untreated control, although this level was still less than what was seen in LPA-treated cells (Fig. 1A-B). To identify the underlying mechanism by which these cells manifested their MT stability, the authors also used C3 toxin (a RhoA inhibitor), which resulted in dramatically reduced levels of Glu MTs (Fig. 1C-D). Using siRNA targeting the downstream effector mDia1, the authors also showed that the effect of Aβ on MT stability is lost under conditions of mDia1 knockdown (Fig. 1E-F).

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This is particularly interesting, as it suggests that Aβ may act to stabilize MTs, and that this may in fact contribute to AD neuropathology. This is surprising in that it seems to counter previous studies that suggest that Aβ acts indirectly to destabilize MTs via tau pathology, although the authors of this research suggest that perhaps these two models are not mutually exclusive; they instead suggest that perhaps the MT destabilization that occurs in the presence of tau may be a compensatory mechanism to counteract the overstabilization of MTs by Aβ. They support this model by adding the fact that hyperphosphorylated tau proteins tend to preferentially target stable Glu MTs that are detyrosinated (Yoshiyama et al., 2003; Rapoport et al., 2002). Ultimately, this finding opens a door to studying the role of Aβ on microtubule function; perhaps, tau pathologies can be pinned to the actions of Aβ on MT dynamics.

Pianu, B., Lefort, R., Thuiliere, L., Tabourier, E., and Bartolini, F. (2014). The A 𝛽1-42 peptide regulates microtubule stability independently of tau. J of Cell Sci. 127(5): 1117–1127.

Karran E, Mercken M, De Strooper B.(2011). The amyloid cascade hypothesis for Alzheimer’s disease: an appraisal for the development of therapeutics. Nat Rev Drug Discov  10(9):698–712. doi:10.1038/nrd3505

Chesarone, M. A., DuPage, A. G. and Goode, B. L. (2010). Unleashing formins to remodel the actin and microtubule cytoskeletons. Nat. Rev. Mol. Cell Biol. 11, 62-74.

Yoshiyama, Y., Zhang, B., Bruce, J., Trojanowski, J. Q. and Lee, V. M. (2003). Reduction of detyrosinated microtubules and Golgi fragmentation are linked to tau-induced degeneration in astrocytes. J. Neurosci. 23, 10662-10671.

To learn more about the work conducted in Dr. Bartolini’s lab, please attend her talk this Tuesday, April 24th at 4:00pm in the Center for Neural Circuits and Behavior.

Junmi Saikia is a third year MD/PhD student who has developed a dangerous addiction to caffeine whilst pursuing research regarding synaptic plasticity and Alzheimer’s disease.

Help, Help I’m being repressed: How repression leads to neuron morphological diversity

One of the most striking features of neurons are their highly diverse dendritic arbors. Neurons can form beautifully complex dendritic arbors allowing different neuron types to have distinct connectivity patterns and functions. Understanding how this large diversity of neuron morphology arises seems like a daunting task. Specific transcription factors have been shown to play prominent roles in generating the massive amount of neuronal diversification observed in the brain. Most of the regulators, however, have been studied separately. How the interactions of these different transcriptional regulators results in neural diversity remains to be understood.

Dr. Wesley Grueber from Columbia University is interested in studying the underlying genetic regulation of neuronal dendritic morphology. Using Drosophila as a model organism, Dr. Grueber’s lab studies how transcriptional regulators can diversify neuronal dendritic arbors. In a recent paper, “Dendritic diversification through transcription factor-mediated suppression of alternative morphologies” Dr. Grueber explores transcriptional strategies for diversifying functionally and morphologically distinct somatosensory neurons in Drosophila.

Drosophila sensory neurons are generally categorized into 4 subtypes based on their dendrite branching patterns and axon targeting. Class I sensory neurons function as proprioceptors, and Class II and III function as touch and gentle touch sensors respectively. Class IV neurons function as multimodal nociceptors. Transcriptional regulators such as Cut have been implicated in promoting the large, complex branching patterns observed in Class II-IV neurons, but is not expressed at detectable levels in neurons with less complex patterns such as Class I neurons. Performing loss-of-function and gain-of-function experiments, Grueber’s lab was able to show that the diversity of the dendritic arbor morphologies emerged from a repressive strategy involving transcription factors Cut, Pmd 1/2, Scalloped, and Vestigial (see fig. 8).


The findings presented in Dr. Grueber’s paper show that repressive transcriptional regulator interactions can generate dendritic diversity. By repressing other transcription factors and target genes, transcription factors can suppress alternative differentiation programs and promote diversity. The authors speculate that transcription factor-mediated suppression of “default programs” in a subset of neurons could be a mechanism by which the diversity of sensory cells types evolved, while maintaining existing modalities.

To hear more about Dr. Wesley Grueber’s research, please attend his talk on Tuesday, April 11, 2017 at 4pm in the CNCB Marilyn G. Farquhar Seminar Room.

Oscar Gonzalez is a first-year graduate student in the neurosciences graduate program and a member of Dr. Maxim Bazhenov’s lab. He is interested in the mechanisms leading to hypersynchronous activity in the brain, and the origin of resting state infra-slow fluctuations.

Cognition and Action: Who’s the Boss?

Would you be happy in a job where every single deliberate and conscious action you made had to be previously approved by your boss? Thankfully, in this scenario, the only one you must answer to is within your brain. The prefrontal cortex (PFC) has long been thought of as the executive control center of goal-directed behavior, and like how a world leader needs to be prepared and able to quickly change strategies as demands and circumstances change within their nation, your PFC facilitates and executes behavior in a fluid, information-sensitive manner.

Research unveiling the mystery of how these executive processes occur have been largely aided by technological advances in neurosurgery. For example, scientists can now implant recording devices directly onto, and sometimes through, the brain surface of patients treated for neurological disorders (e.g. epilepsy), unveiling a wealth of data that is collected as the patient is awake and performing various tasks. More importantly, these intracranial recordings (electrocorticography; ECoG) are highly precise in their spatial and temporal resolution, providing an arguably better idea of brain function compared to other prominent imaging techniques, such as the less temporally precise functional magnetic resonance imaging method. In ECoG, tiny electrodes record local field potentials (LFPs), which serve as a proxy for the summed electric current flowing from a collection of neurons.


Intracranial electrode grid for electrocorticography (ECoG) [Wikipedia Commons]

How decision making occurs has begun to be looked at an elevated level compared to single neuron activity; while the individual activation of single brain cells can be highly associated with cognitive phenomena, their spiking and excitability properties can be better thought of as being a consequence sustained activation or interactions at the population level. For instance, human intracranial data filtered to look at high-frequency (70-200 Hz) activity revealed that the PFC became active only when unpredicted deviants (e.g. errors) were detected in an auditory attention task. At Robert Knight’s cognitive neuroscience research laboratory at the University of California, Berkeley, it has recently been proposed that such oscillatory dynamics at the network level reflect “activity-silent” encoding of rules relevant to the task at hand. For example, while active processing by the brain’s executive control center has been thought of as increase in neuronal spiking or high frequency activity, cognitive phenomena, such as prediction error detection, has been observed to arise without changes in individual spiking activity. This “activity silent” encoding is thought to reflect endogenous oscillatory activity that may underlie how PFC guides behavior.


“Different contexts could be embedded in distinct spatiotemporal configurations of the same network” [Randolph & Knight, 2016]

Come check out Dr. Knight’s talk, titled “Insights into human cognition from intracranial recording”, on Tuesday, April 4th, at 4 P.M. in the CNCB Marilyn Farquhar Seminar Room.

Christian Cazares is a first-year neuroscience graduate student rotating in the Chalasani Lab. He can be reached at @fleabrained and

Randolph F. H. and Knight R. T. (2016) Oscillatory Dynamics of Prefrontal Cognitive Control Trends in Cognitive Sciences, Volume 20 , Issue 12 , 916 – 930 staff (2014). “Medical gallery of Blausen Medical 2014”. WikiJournal of Medicine 1 (2). DOI:10.15347/wjm/2014.010. ISSN 2002-4436.

Oh, to regenerate axons! Hypoxia inducible factor’s role in axonal regeneration

How does the nervous system respond to axonal injury? The answer is: it depends. In the environment of the central nervous system (CNS), neurons possess extremely low regenerative capacity. In contrast, the peripheral nervous system’s (PNS) neurons exhibit the ability to regrow following injury. While the mechanisms underlying this regrowth is not completely understood, there is significant motivation to elucidate this field for the purpose of developing therapies to enhance axonal regeneration even in the inhibitory environment of the CNS.

Dr. Valeria Cavalli’s group at Washington University decided to study axonal regeneration in the context of the dorsal root ganglion (DRG). What makes the DRG neuron an interesting model system is its structure as a pseudounipolar neuron: from the cell bodies located in the dorsal root ganglion located adjacent to the spinal cord arises a single extension which splits into two branches: a central branch that navigates back to the spinal cord, and a peripheral branch, which as the name suggests projects peripherally. Injury to this peripheral branch of the DRG results in activation of a pro-regenerative program – that is, a series of molecular events that promote the regrowth of the damaged axon. The central branch, keeping in theme with the CNS, does not exhibit such regenerative capacities. What, then, allows a single DRG neuron to exhibit repair on one branch but not the other?

Transcriptional profiling studies in DRG neurons identified a handful of interesting genes upregulated following injury; among these is the gene known as Hypoxia Inducible Factor (HIF), a heterodimeric protein made of HIF-α and HIF-β subunits. HIF-α has a particularly interesting expression profile: at physiologic O2 concentrations, HIF-α is targeted for ubiquitination/degradation pathways. As O2 concentrations drop, this suppression of HIF-α expression is released.

Cho et al., 2015, showed that following peripheral DRG axotomy, a large percentage of HIF-α targeted genes are upregulated over a 1.2 fold threshold; furthermore, this upregulation of gene expression is lost in HIF-α knockdown DRGs. In uninjured DRG neurons, constitutive overexpression of HIF-α, but not knockdown of HIF-α, lead to significant changes in gene expression compared to wildtype uninjured DRG neurons.

To study the regenerative capacity of DRG neurons, Cho et al. infected DRG neurons either with control shRNA, two HIF-α targeting shRNAs, or a lentivirus virally overexpresses HIF-α. These neurons were axotomized and immunostained for SCG10, a marker of axon regeneration. Interestingly, both shRNA-mediated knockdown and lentiviral overexpression of constitutively active HIF-α resulted in reduced SCG10 fluorescence, suggesting that regulated expression of HIF-α (as opposed to constitutive overexpression) is necessary for axonal regrowth. Further downstream, Cho et al. conducted knockdowns of genes targeted by HIF-α, and showed that while knockdowns contributed to reduced axon regeneration for some genes, there were still others genes that, when knocked down, instead exhibited slight increases in axon regeneration. This would suggest that HIF-α targeted genes could either promote or inhibit axon repair.

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In order to show how HIF-α regulates axon regrowth in vivo, Cho et al. generated mice lacking HIF-α in sensory neurons by crossing HIFAflox/flox with mice expressing Cre under the Advillin promoter (HIF1AcKO), which is specific to the peripheral sensory nervous system. After confirming HIF-α knockout, the experimenters crushed the sciatic nerve and quantified regeneration of the nerve axons 3 days later by SCG10 staining. Using SCG10 intensity and distance from crush site, the researchers calculated a regeneration index; in the HIF1AcKO, regeneration was limited past the crush site (Fig 4A-C). Injury to the sciatic nerve, but not the dorsal root itself, results in elevated HIF-α expression (Fig 4D,E).

While hypoxia does increase HIF-α levels in vitro, Cho et. al specifically showed that acute intermittent hypoxia (AIH) enhances axon regeneration in vivo via elevating HIF-α and subsequent downstream target genes such as vascular endothelial growth factor (VEGFA). They demonstrated axon regeneration both in the sensory sciatic nerve fibers as well as in sciatic motor neuron: sciatic injury in YFP-16 mice, which express yellow fluorescent protein (YFP) in motor neurons, showed increased colocalization of YFP and neuromuscular junction boutons following AIH treatment for sciatic nerve injury. Taken together, these results suggests that AIH may drive axon regeneration in the periphery via HIF-α-dependent mechanisms.

To learn more about the work conducted in Dr. Cavalli’s lab, attend her talk this Tuesday, March 21th, at 4:00pm in the Center for Neural Circuits and Behavior.

Junmi Saikia is a third year MD/PhD student in the Malinow Lab at UCSD. Her research interests revolve around neurodegenerative diseases.  She apologizes in advance  for the punishably punfunny title.

Ripples in time: How the hippocampus helps us learn, not just during sleep

We’ve all at some point been told that sleep is essential for our memories to be consolidated: when we sleep, our hippocampus takes all of the things we’ve learned during the day and files them away for easy access when we’ll need them next. Known contributors to this phenomenon are Sharp-wave-ripple events, or SWRs, are a brief (lasting only 100-200ms) oscillation of hippocampal neural activity occurring at approximately 200Hz. Interestingly, they occur not only during slow-wave sleep, but also during awake stillness—and although their presence during sleep has frequently been attributed to the memory consolidation process, their function during awake states is unclear. What role might SWRs play while we’re awake, and how does their occurrence affect our behavior?

Dr. A. David Redish at the University of Minnesota is interested in precisely these questions. His lab combines the use of multi-electrode recordings with computational analysis that allow for the study of neural ensemble activity at high temporal resolution. A recent (2016) paper from Dr. Redish’s lab, titled “Interplay between Hippocampal Sharp-Wave-Ripple Events and Vicarious Trial and Error Behaviors in Decision Making”, elucidates a possible role for sharp-wave-ripple events in the context of decision-making.

Redish and his group quantified the presence of Vicarious Trial and Error (VTE) behaviors—a measure of uncertainty and deliberation—in relation to the presence of SWEs captured through neural recordings as rats completed several complex navigation tasks to obtain rewards. These behavioral tasks enabled the experimenters to measure the rats’ learning, deliberating, and decision-making in the context of SWE occurrences. The experimenters were ultimately able to observe an association between SWEs in awake states and VTE behaviors as rats decided which path to take in the tasks.

Screen Shot 2017-03-13 at 9.42.16 AM

Part A in the figure above demonstrates the nature of the spatial alternation task. A1 and A2 comprise the first trial, in which the rat had to exit the center arm and move to a side arm to obtain a reward, then return to its starting position in the center arm to obtain a second reward. The second trial, in A3 and A4, shows that the rat needed to travel to the opposite side arm from the first trial (and back to center) in order to obtain the rewards. Part B illustrates a key finding of Dr. Redish’s paper: that the disruption of hippocampal sharp-wave-ripple events (SWR) increased the proportion of VTE behaviors.

The results of Dr. Redish’s study suggest that SWRs and VTE behaviors are inversely correlated. Moreover, because VTE behaviors are associated with increased behavioral uncertainty and variability, they imply that SWRs and associated mechanisms engaged in learning and memory might play a specific role in decision-making once a particular behavior or reward has been learned. Thus, there is clearly a complex interplay of these two processes occurring during awake learning and navigation that demands exploration in greater detail.

To learn more about the work conducted in Dr. Redish’s lab, attend his talk this Tuesday, March 7th, at 4:00pm in the Center for Neural Circuits and Behavior.

Marley Rossa is a first-year Ph.D. student whose research interests encompass the use of computational techniques to analyze neural data and thereby understand the nature of information communicated between neurons and neural ensembles.



Divide and Conquer! How the Brain Avoids Sensory Overload

They say that he human brain is the most complex machine in the universe, and it’s for good reason. Brains are comprised of about a hundred billion neurons that typically form a thousand unique connections each. As a result scientists estimate that the brain can store ten to a thousand times more information than modern day laptops.

computer-brain-interface-darpa-sHowever, what is even more impressive than the storage capacity of the brain is its ability to dynamically read and process massive amounts of information. Every second that you are awake, your brain is efficiently parsing through constantly flowing streams of visual, auditory, gustatory, and olfactory data. How does the brain sort through all this rich information? What mechanisms underlie our ability to prioritize important sensory information when presented with competing or conflicting cues?

These open questions in neuroscience have long puzzled scientists. In this post, we will take a look at the paper from Michael Halassa’s group, titled “Thalamic control of sensory selection in divided attention”,  that sheds some light on these questions. The central goal of the article is to provide some proof for the hypothesis that the frontal cortex, the home of executive function, regulates the selection of sensory information in the thalamus, the hub of sensory inputs into the brain.

To accomplish this, Halassa first creates a rodent psychometric test that basically tests the ability of a mouse to secure a food reward when presented with either congruent or conflicting visual and auditory stimuli.  As expected, mice were forced to divide their attention when presented with both visual and auditory information. When presented with conflicting auditory tones, mice had a higher visual detection threshold, which persisted even as the tones were switched back to being congruent. This provided evidence that top-down processing, likely from the frontal cortex was biasing the visual stream! Next the authors sought to pin down the circuit mechanisms of this finding.

Next using optogenetics, a technique to make neurons respond to light, Halassa’s group linked channelrhodopsin to GABAergic inhibitory cells in the frontal cortex. This allowed them to effectively silence activity between a region of the frontal cortex that sends projections to the thalamus, the sensory hub. In the same behavioral tests, the mice perform much worse when the link between the frontal cortex and the thalamus is inhibited. Without this connection, mice are suddenly unable to sort out conflicting sensory cues in order to complete the task!screen-shot-2017-03-06-at-1-30-21-am

Importantly, the paper also shows that a similar sort of disruption scheme in the visual cortex fails to disrupt task performance in a similar manner. This means that the cross-cortical interactions are less important than corticothalamic connections, which was previously not known.

In order to fully characterize the circuit, optogenetic methods were combined with viral tracing to follow the projections from the frontal cortex to a particular region in the thalamus, called the thalamic reticular nucleus (RTN). Using electrophysiology and optogenetics, it is shown the neurons in this region fire specifically when conflicting auditory tones are played, and that silencing this region results in the a similar hit in task performance, as above. Quite convincingly, it seems the the thalamic target of the frontal cortex for sensory selection is the RTN.


To wrap this study up, the authors investigated the interaction between the RTN and adjacent lateral geniculate nucleus (LGN), a thalamic structure that is crucial for visual processing. Using a novel assay for in vivo inhibitory activity (FRET photometry), they demonstrate that when mice are attending to conflicting auditory cues, the RTN uses direct feed-forward inhibition, as measured by intracellular chloride ion shifts, on the LGN to alter visual perception.

In summary, this article elegantly describes the link between the frontal cortex, the thalamus, and the visual system using a plethora of high-powered and novel experimental techniques. The authors provide convincing evidence for a mechanism in which the more “intelligent” regions of the brain regulate the flow of sensory information such that cues relevant to the task at hand are amplified and useless cues are discarded.

To learn more about the work conducted in Dr. Halassa’s lab, please attend his talk this Tuesday, March 7th, at 4:00pm in the Center for Neural Circuits and Behavior.

Debha Amatya is a third year MD/PhD student in the Laboratory of Genetics at the Salk Institute. He’s interested in psychiatric genomics and weightlifting. 

Optogenetic regulation of dopamine neurons in the midbrain regulates depression-related behaviors in mice

In 2015, 4% of U.S. adults aged 18 or older experienced a serious mental illness that interfered with daily functioning, and this percentage increases to almost 18% when adults with any mental illness are considered. Major depression in particular is one of the most common psychiatric disorders in the U.S., and has the highest disability burden among all mental disorders. Despite the social and economic costs of depression, relatively little is known about its neurobiological basis.

Anhedonia, or the inability to experience pleasure from activities usually found to be enjoyable, is a key feature of depression, and suggests that there may be a deficit in the reward circuitry of the brain in this disorder. Dopaminergic neurons in the ventral tegmental area (VTA) of the midbrain that project to the nucleus accumbens (NAc) have been implicated in reward recognition and initiation of reward utilization. Alterations in the function of this pathway could potentially underlie some of the behavioral deficits seen in people with depression.

Two in vivo firing patterns have been identified in dopaminergic VTA neurons: 1) low frequency tonic firing, and 2) high frequency phasic firing. Previously, phasic firing was shown to be upregulated by repeated social-defeat stress (a mouse model of depression) in susceptible mice, whereas mice resilient to the effects of repeated social-defeat showed no firing rate change.

Dr. Eric Nestler’s lab at Icahn School of Medicine at Mount Sinai in New York used optogenetic techniques to drive different firing patterns in dopaminergic VTA neurons in order to assess whether phasic firing patterns could induce depression-like behaviors in mice. They injected double floxed Cre-dependent AAV virus expressing channelrhodopsin-2 fused with enhanced yellow fluorescent protein (ChR2-eYFP) into the VTA of tyrosine hydroxylase (TH)-Cre mice to target expression of ChR2-eYFP to dopaminergic VTA neurons. VTA neurons were driven via activation of ChR2 at either a low frequency tonic pattern or a high frequency phasic pattern during a subthreshold social-defeat stress paradigm. Mice that received phasic stimulation showed a significant increase in depression-like behavior compared to mice receiving tonic stimulation. The depression-like phenotype was characterized by decreased social interaction with another mouse and decreased sucrose preference.


Chaudhury et al. (2013) then assessed whether phasic firing in VTA neurons could confer susceptibility in mice resilient to the social-defeat paradigm. A standard 10-day social-defeat paradigm was performed in wild-type mice, and a social-interaction test was then performed to identify resilient mice. HSV-ChR2-eYFP virus was injected into the VTA of resilient mice, and a second social-interaction test was then performed while undergoing optogenetic phasic stimulation of VTA neurons. The behavioral phenotype of these mice switched from resilient to susceptible, as shown by a decrease in social interaction and a decrease in sucrose preference 12 hours after optogenetic activation of VTA neurons. VTA neurons exhibited sustained alterations in excitability 12 hours after optogenetic activation, evidenced by increased spontaneous and evoked activity. This increased excitability may explain the decreased sucrose preference seen 12 hours after VTA neuronal stimulation in the social-interaction test.

Given that VTA dopamine neurons project to numerous brain areas, they wanted to test the role of the VTA-NAc pathway specifically in mediating the depression-related phenotype. Replication-defective pseudorabies virus expressing Cre (PRV-Cre) was injected into the NAc, and the Cre-dependent AAV ChR2-eYFP virus was injected into the VTA. This combination caused expression of ChR2 in only NAc-projecting VTA neurons. Mice were subjected to the subthreshold social-defeat paradigm, and then 24 hours later optogenetic stimulation of VTA neurons was performed during the social-interaction test. This manipulation reliably induced the depression-related phenotype of increased social avoidance and decreased sucrose preference. Similar methodology was employed when testing the effects of inhibiting this pathway, but an AAV halorhodopsin-eYFP virus was injected into the VTA instead. Inhibition of VTA neurons during the social-interaction test induced resilience to the depression-related phenotype, with increased social interaction and increased sucrose preference.


Manipulation of firing rates in VTA neurons projecting to the medial prefrontal cortex (mPFC) were also performed in this study using ChR2 and halorhodopsin, and the same methodologies that were used to study the VTA-NAc pathways were employed. Optogenetically induced phasic firing of VTA-mPFC neurons had no effect on social interaction and sucrose preference after subthreshold social defeat. Inhibition of VTA-mPFC neuronal activity via halorhodopsin activation during the social-interaction test induced a susceptible phenotype (decreased social interaction) after subthreshold social defeat, but no difference in sucrose preference was observed.

The results of these studies delineate neural circuitry involved in the induction of depression-like behavior after social-defeat stress, specifically the VTA-NAc pathway mediating anhedonia-like behavior. As we begin to unravel the neurobiological complexity of depression and other psychiatric illnesses, new therapeutic strategies can be created to more selectively target pathways responsible for the behavioral abnormalities seen in these disorders.

To learn more about the work conducted in Dr. Nestler’s lab, please attend his talk this Tuesday, February 28, at 4:00pm in the Center for Neural Circuits and Behavior.

Molly Kwiatkowski is a third year MD/PhD student in the Young lab located in the Consortium for Translational Research in Neuropsychopharmacology. Her work focuses on elucidating neural mechanisms underlying psychiatric disorders such as bipolar disorder.


Analyzing Associations: Differential Amygdalar Activity to Conditioned Stimuli

Associative learning, the process by which an organism forms a link between a cue or behavior and an intrinsically valenced stimulus, is a critical feature of adaptive behavior and survival. Learning to run upon hearing a roar associated with a predator allows one to avoid getting maimed or eaten; learning to rush to the kitchen when you smell freshly baked cookies enables you to snag a warm, tasty treat. As demonstrated by these two examples, associations can occur with both negatively valenced and positively valenced stimuli, with the former often invoking withdrawal and avoidance behaviors and the latter enforcing active approaching behaviors. Because associative memory acts as such a cornerstone in shaping flexible and appropriate behaviors, when this process goes awry, various negative consequences can ensue: depression, substance abuse, and anxiety disorder are only a few of such maladies tied to disturbances in circuitry underlying associative and motivational functionality. Consequently, studying this subject is of paramount therapeutic, as well as intellectual, interest.

Dr. Kay Tye’s Lab at the Massachusetts Institute of Technology aims to understand these very processes, employing a variety of methods from optogenetics and electrophysiological recordings to pharmacological and imaging techniques. A recent paper published by Dr. Tye and colleagues provides significant insight into the routing of positive and negative information from the amygdala during memory retrieval, suggesting that specific cell populations in the basolateral amygdala (BLA) encode different types of emotional valence depending upon the region to which they project. While the BLA as a whole is crucial in forming both positively and negatively valenced associations, subpopulations within the BLA demonstrate distinct activity patterns to rewarding versus aversive conditioned stimuli.

The main targets of the BLA investigated in Beyeler et al.’s (2016) study included the nucleus accumbens (NAc), central amygdala (CeA), and ventral hippocampus (vHPC). While the NAc is generally involved in positive reinforcement, the CeA is linked to learning that mediates aversive behaviors, and the vHPC presumably promotes anxiogenic actions. To examine the neural codes of BLA neurons targeting the NAc, CeA, and vHPC, Beyeler et al. (2016) utilized a dual-virus approach entailing optogenetic-mediated phototagging and in vivo electrophysiological recordings in mice trained to associate a particular tone with sucrose (a rewarding outcome) delivery and another tone with quinine delivery (an aversive outcome). After the mice learned the associations between each stimulus and its respective outcome—assessed as showing anticipatory licking selectively after hearing the sucrose-predictive tone for 70% of the trials—Beyeler et al. (2016) performed acute recordings in the BLA.

Half of the neurons recorded in the BLA were found to respond to sucrose and/or quinine, with 28% of neurons demonstrating selective responses to the tone associated with sucrose and a mere 9% altering firing selectively to the tone associated with quinine. In addition, 13% of BLA neurons responded to both tones similarly, and less than 1% responded to both tones in qualitatively different manners.



Nevertheless, the most notable finding relates to the analysis of the subpopulations in the BLA that responded to both or to either of these auditory cues. Specifically, the subpopulation of BLA neurons that projected to the NAc exhibited a starkly different response profile than that of the BLA-CeA neurons. Zero BLA-NAc neurons were found to be excited by the tone associated with quinine, but nearly all (77%) were excited by the tone tied to sucrose; Any BLA-NAc neurons that reacted to the quinine-associated tone demonstrated inhibition by the conditioned stimulus. In contrast, all BLA-CeA projectors recorded were excited by the tone paired with quinine.

Classifying BLA projector populations using “valence bias”—considering excitation and inhibition as qualitatively different—reinforced the differential activity of the three subpopulations examined. Approximately half of the BLA population encoding valence showed a positive bias—increasing firing for the sucrose-associated tone and/or decreasing activity to the quinine-associated tone—while the other half appeared to encode negative valence—increasing firing to the quinine cue and decreasing firing to the sucrose cue. Within the BLA, a greater percentage of BLA-NAc projectors were observed to be encode positive valence (80%, compared to 31% in the BLA-CeA); whereas more BLA-CeA cells encoded negative valence (69%). Interestingly, the BLA-vHPC subpopulation responded evenly to both tones, with 43% encoding positive valence.


These findings clearly suggest that BLA-NAc neurons preferentially encode cues associated with rewarding outcomes, consistent with the known role of the NAc in positive reinforcement; and that BLA-CeA neurons preferentially encode cues linked to aversive outcomes, also consistent with its acknowledged role in assigning negative valence to stimuli. The observation of BLA-vHPC responses evenly distributed between positive and negative valence is somewhat surprising due to its supposed role in anxiogenic behavior and requires further investigation.

If you find this experiment intriguing and would like to learn more about the nuances of and mechanisms underlying neural systems supporting associative learning, motivation,  and emotional processing, attend Dr. Kay Tye’s talk this Tuesday, February 21, at 4:00 pm at the Center for Neural Circuits and Behavior.


Gina D’Andrea-Penna is a first-year neurosciences Ph.D. student whose interests largely reside in the domain of cognitive neuroscience, particularly in investigating perceptual awareness, attention, and working memory.