Biology subjects

Nelson, S. B.

Publications and source records attributed to Nelson, S. B..

11 recordsLinked to original sources

bioRxiv2025

Multiplexed CRISPRi Reveals a Transcriptional Switch Between KLF Activators and Repressors in the Maturing Neocortex

A critical phase of mammalian brain development takes place after birth. Neurons of the mouse neocortex undergo dramatic changes in their morphology, physiology, and synaptic connections during the first postnatal month, while properties of immature neurons, such as the capacity for robust axon outgrowth, are lost. The genetic and epigenetic programs controlling prenatal development are well studied, but our understanding of the transcriptional mechanisms that regulate postnatal neuronal maturation is comparatively lacking. By integrating chromatin accessibility and gene expression data from two subtypes of neocortical pyramidal neurons in the neonatal and maturing brain, we predicted a role for the Kruppel-Like Factor (KLF) family of Transcription Factors in the developmental regulation of neonatally expressed genes. Using a multiplexed CRISPR Interference (CRISPRi) knockdown strategy, we found that a shift in expression from KLF activators (Klf6, Klf7) to repressors (Klf9, Klf13) during early postnatal development functions as a transcriptional switch to first activate, then repress a set of shared targets with cytoskeletal functions including Tubb2b and Dpysl3. We demonstrate that this switch is buffered by redundancy between KLF paralogs, which our multiplexed CRISPRi strategy is equipped to overcome and study. Our results indicate that competition between activators and repressors within the KLF family regulates a conserved component of the postnatal maturation program that may underlie the loss of intrinsic axon growth in maturing neurons. This could facilitate the transition from axon growth to synaptic refinement required to stabilize mature circuits.

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bioRxiv2023

Progressive circuit hyperexcitability in mouse neocortical slice cultures with increasing duration of activity silencing

Forebrain neurons deprived of activity become hyperactive when activity is restored. Rebound activity has been linked to spontaneous seizures in vivo following prolonged activity blockade. Here we measured the time course of rebound activity and the contributing circuit mechanisms using calcium imaging, synaptic staining and whole cell patch clamp in organotypic slice cultures of mouse neocortex. Calcium imaging revealed hypersynchronous activity increasing in intensity with longer periods of deprivation. While activity partially recovered three days after slices were released from five days of deprivation, they were less able to recover after ten days of deprivation. However, even after the longer period of deprivation, activity patterns eventually returned to baseline levels. The degree of deprivation-induced rebound was age-dependent, with the greatest effects occurring when silencing began in the second week. Pharmacological blockade of NMDA receptors indicated that hypersynchronous rebound activity did not require Hebbian plasticity evoked. In single neuron recordings, input resistance roughly doubled with a concomitant increase in intrinsic excitability. Synaptic imaging of pre- and postsynaptic proteins revealed dramatic reductions in the number of presumptive synapses with a larger effect on inhibitory than seen in excitatory synapses. Putative excitatory synapses colocalizing PSD-95 and Bassoon declined by 39% and 56% following five and ten days of deprivation, but presumptive inhibitory synapses colocalizing gephyrin and VGAT declined by 55% and 73% respectively. The results suggest that with prolonged deprivation, a progressive reduction in synapse number is accompanied by a shift in the balance between excitation and inhibition and increased cellular excitability. SIGNIFICANCE STATEMENTWhen cortical activity is silenced during development, lifelong seizures often result. Here we explored whether these seizures result from overcompensation of homeostatic recovery mechanisms. Prior work showed that neurons briefly deprived of the ability to fire action potentials compensate by becoming more excitable, increasing synaptic drive and intrinsic excitability. We found that prolonged silencing of cortex produced a profound loss of synapse density, especially for inhibitory synapses, pointing to a circuit unable to maintain excitatory/inhibitory balance. These results show that homeostatic responses, which are normally restorative, can result in maladaptive circuit configurations when brought to extremes.

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bioRxiv2023

Prolonged activity-deprivation causes pre- and postsynaptic compensatory plasticity at neocortical excitatory synapses

Homeostatic plasticity stabilizes firing rates of neurons, but the pressure to restore low activity rates can significantly alter synaptic and cellular properties. Most previous studies of homeostatic readjustment to complete activity silencing in rodent forebrain have examined changes after two days of deprivation, but it is known that longer periods of deprivation can produce adverse effects. To better understand the mechanisms underlying these effects and to address how presynaptic as well as postsynaptic compartments change during homeostatic plasticity, we subjected mouse cortical slice cultures to a more severe five-day deprivation paradigm. We developed and validated a computational framework to measure the number and morphology of presynaptic and postsynaptic compartments from super resolution light microscopy images of dense cortical tissue. Using these tools, combined with electrophysiological miniature excitatory postsynaptic current measurements, and synaptic imaging at the electron microscopy level, we assessed the functional and morphological results of prolonged deprivation. Excitatory synapses were strengthened both presynaptically and postsynaptically. Surprisingly, we also observed a decrement in the density of excitatory synapses, both as measured from colocalized staining of pre- and postsynaptic proteins in tissue, and from the number of dendritic spines. Overall, our results suggest that cortical networks deprived of activity progressively move towards a smaller population of stronger synapses. SIGNIFICANCE STATEMENTBlocking activity in neocortical slice cultures produced coordinated pre and postsynaptic changes at excitatory synapses. Functional and structural assays suggest that deprivation results in fewer excitatory synapses, but each is strengthened both pre- and postsynaptically. This may contribute to the emergence of epileptiform activity.

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bioRxiv2021

A transcriptional constraint mechanism limits the homeostatic response to activity deprivation in mammalian neocortex

Healthy neuronal networks rely on homeostatic plasticity to maintain stable firing rates despite changing synaptic drive. These mechanisms, however, can themselves be destabilizing if activated inappropriately or excessively. For example, prolonged activity deprivation can lead to rebound hyperactivity and seizures. While many forms of homeostasis have been described, whether and how the magnitude of homeostatic plasticity is constrained remains unknown. Here we uncover negative regulation of cortical network homeostasis by the PARbZIP family of transcription factors. In cortical slice cultures made from knockout mice lacking all three of these factors, the network response to prolonged activity withdrawal measured with calcium imaging is much stronger, while baseline activity is unchanged. Whole cell recordings reveal an exaggerated increase in the frequency of miniature excitatory synaptic currents reflecting enhanced upregulation of recurrent excitatory synaptic transmission. Genetic analyses reveal that two of the factors, Hlf and Tef, are critical for constraining plasticity and for preventing life-threatening seizures. These data indicate that transcriptional activation is not only required for many forms of homeostatic plasticity but is also involved in restraint of the response to activity deprivation.

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bioRxiv2021

Loss of Stk11 in basolateral amygdalar projection neurons impairs CTA learning by altering the temporal pattern of taste response plasticity in GC

Gustatory cortex (GC) responds to tastes on the tongue with dynamic ensemble activity that represents first the presence, then the identity, and finally the hedonic value (palatability) of tastes. This final state of the taste response is uniquely altered by conditioned taste aversion (CTA) - a powerful one-trial learning paradigm in which a taste becomes aversive after association with gastric malaise - a process requiring coordination between GC and basolateral amygdala (BLA). Previously, we found that one key requirement for learning in this circuit is expression of the serine/threonine kinase 11 (Stk11) gene (a tumor suppression gene which has only recently been associated with learning; Levitan et al., 2020). When Stk11 is knocked out in BLA projection neurons (BLApn), CTA learning fails to occur. Here we have examined how learning-related response plasticity in GC taste responses is impacted by the knockout of Stk11 in BLApn. Contrary to the commonly held assumption that a lack of learning means a lack of such plasticity, but consistent with the fact that Stk11KO has been shown to increase the excitability of BLApn, our data reveal that the knockout of Stk11 in BLApn does not eliminate plasticity; rather, it shifts the impact of CTA training on GC taste responses to an earlier, learning-inappropriate epoch. Even naive taste representations are altered--specifically, the pattern of similarities and differences among the different taste responses are rendered abnormal by Stk11 KO, and these relationships fail to change with training. Finally, the latency of behavior-related dynamic ensemble features of the GC taste response, which is also abnormal in naive KO mice, is rendered disorganized by CTA. Together, these results suggest that Stk11 plays a role in governing the coordination of GC activity by BLA, and demonstrate that alterations in the function of BLApn caused by Stk11KO inhibit learning not by inhibiting plasticity but by changing its temporal properties.

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bioRxiv2019

Deletion of Stk11 and Fos in BLA projection neurons alters their intrinsic excitability and impairs formation of long-term aversive memory

Conditioned taste aversion (CTA) is a form of one-trial learning dependent on basolateral amygdala projection neurons (BLApn). Its underlying cellular and molecular mechanisms are poorly understood, however. We used RNAseq from BLApn to identify learning-related changes in Stk11, a kinase with well-studied roles in growth, metabolism and development, but not previously implicated in learning. Deletion of Stk11 restricted to BLApn completely blocks memory when occurring prior to training, but not following it, despite altering neither BLApn-dependent encoding of taste palatability in gustatory cortex, nor transcriptional activation of BLApn during training. Deletion of Stk11 in BLApn also increases their intrinsic excitability. Conversely, BLApn activated by CTA to express the immediate early gene Fos had reduced excitability. BLApn knockout of Fos also increased excitability and impaired learning. These data suggest that Stk11 and Fos expression play key roles in CTA long-term memory formation, perhaps by modulating the intrinsic excitability of BLApn.

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bioRxiv2019

Cortical RORβ is required for layer 4 transcriptional identity and barrel integrity.

Retinoic Acid-Related Orphan Receptor Beta (ROR{beta}) is a transcription factor (TF) and marker of layer 4 (L4) neurons, which are distinctive both in transcriptional identity and the ability to form aggregates such as barrels in rodent somatosensory cortex. However, the relationship between transcriptional identity and L4 cytoarchitecture is largely unknown. We find ROR{beta} is required in the cortex for L4 aggregation into barrels and thalamocortical afferent (TCA) segregation. Interestingly, barrel organization also degrades with age. Loss of ROR{beta} delays excitatory input and disrupts gene expression and chromatin accessibility, with downregulation of L4 and upregulation of L5 genes, suggesting a shift in cellular identity. Expression and binding site accessibility change for many other TFs, including closure of neurodevelopmental TF binding sites and increased expression and binding capacity of activity-regulated TFs. Lastly, a putative target of ROR{beta}, Thsd7a, is downregulated without ROR{beta}, and Thsd7a knockout alone disrupts TCA organization in adult barrels.

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bioRxiv2019

Single and population coding of taste in the gustatory cortex of awake mice

Electrophysiological analysis has reveals much about the broad coding and neural ensemble dynamics that characterize gustatory cortical (GC) taste processing in awake rats, and about how these dynamics relate to behavior. With regard to mice, meanwhile, data concerning cortical taste coding have largely been restricted to imaging--a technique that reveals average levels of neural responsiveness, but that (currently) lacks the temporal sensitivity necessary for evaluation of fast response dynamics; furthermore, the few extant studies have thus far failed to provide consensus on basic features of coding. We have recorded the spiking activity of ensembles of GC neurons while presenting representatives of the basic taste modalities (sweet, salty, sour and bitter) to awake mice. Our first central result is the identification of similarities between rat and mouse taste processing: most mouse GC neurons (~66%) responded distinctly to multiple (3-4) tastes; temporal coding analyses further reveal, for the first time, that single mouse GC neurons sequentially code taste identity and palatability--the latter responses emerging ~0.5s after the former--with whole GC ensembles transitioning suddenly and coherently from coding taste identity to coding taste palatability. The second finding is that spatial location plays very little role in any aspect of taste responses--neither between- (anterior-posterior) nor within-mouse (dorsal-ventral) mapping revealed anatomical regions with narrow or temporally simple taste responses. These data confirm recent results showing that mouse cortical taste responses are not \"gustatopic,\" but also go beyond these imaging results to show that mice process tastes through time.

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bioRxiv2018

The role of the gustatory cortex in incidental experience-evoked enhancement of later taste learning

The strength of learned associations between pairs of stimuli is affected by multiple factors, the most extensively studied of which is prior experience with the stimuli themselves. In contrast, little data is available regarding how experience with incidental stimuli (independent of any conditioning situation) impacts later learning. This lack of research is striking given the importance of incidental experience to survival. We have recently begun to fill this void using conditioned taste aversion (CTA), wherein an animal learns to avoid a taste that has been associated with malaise. We previously demonstrated that incidental exposure to salty and sour tastes (taste pre-exposure, TPE) enhances aversions learned later to sucrose. Here, we investigate the neurobiology underlying this phenomenon. First, we use immediate early gene (cFos) expression to identify gustatory cortex (GC) as a site at which TPE specifically increases the neural activation caused by taste-malaise pairing (i.e., TPE did not change cFos induced by either stimulus in isolation). Next, we use site-specific infection with the optical silencer Archaerhodopsin-T to show that GC inactivation during TPE inhibits the expected enhancements of both learning and CTA related cFos expression, a full day later. Thus, we conclude that GC is almost certainly a vital part of the circuit that integrates incidental experience into later associative learning.

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bioRxiv2017

A topographic axis of transcriptional identity in thalamus

Uncovering common principles by which diverse modalities of information are processed is a fundamental goal in neuroscience. In mammalian brain, thalamus is the central processing station for inputs from sensory systems, subcortical motor systems, and cortex; a function subserved by over 30 defined nuclei1,2. Multiple thalamic nuclei send convergent information to each region of the forebrain, but whether there is a conserved architecture across the set of thalamic pathways projecting to each forebrain area has remained unresolved3-5. To uncover organizational principles of thalamic pathways, we produced a near-comprehensive transcriptomic atlas of thalamus. This revealed a common logic for thalamic nuclei serving all major cortical modalities. We found that almost all nuclei belong to one of three major profiles, with a given cortical area getting input from each of these profiles. These profiles lie on a single axis of variance aligned with the mediolateral axis of thalamus, and this axis is strongly enriched in genes encoding receptors and ion channels. We further show that each projection profile exhibits different electrophysiological signatures. Single-cell profiling revealed that rather than forming discrete classes, thalamic neurons lie on a spectrum, with intermediate cells existing between profiles. Thus, in contrast to canonical models of thalamus that suggest it is a switchboard primarily concerned with routing distinct modalities of information to distinct cortical regions, we show that the thalamocortical system is more akin to a molecularly-defined filter bank repeatedly applied across modality. Together, we reveal striking covariation in the organization of thalamic pathways serving all input modalities and output targets, establishing a simple and comprehensive thalamic functional architecture.

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bioRxiv2017

The Transcriptional Logic of Mammalian Neuronal Diversity

The mammalian nervous system is constructed of many cell types, but the principles underlying this diversity are poorly understood. To assess brain-wide transcriptional diversity, we sequenced the transcriptomes of the largest collection of genetically and anatomically identified neuronal classes. Using improved expression metrics that distinguish information content from signal-to-noise-ratio, we found that homeobox transcription factors contain the highest information about cell types and have the lowest noise. Genes that contribute the most to neuronal diversity tend to be long and enriched in factors specifically involved in neuronal function. Genome accessibility measurements reveal that long genes have more candidate regulatory elements arrayed in more distinct patterns. These elements frequently overlap interspersed repeats (mobile elements) and the pattern of repeats is predictive of gene expression. New regulatory sites resulting from elongation of neuronal genes by mobile elements may be an evolutionary force enhancing nervous system complexity.

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