Audra Van Wart

Polarized Distribution of Ion Channels within Microdomains of the Axon Initial Segment

Voltage‐gated sodium (Nav) channels accumulate at the axon initial segment (IS), where their high density supports spike initiation. Maintenance of this high density of Nav channels involves a macromolecular complex that includes the cytoskeletal linker protein ankyrin‐G, the only protein known to bind Nav channels and localize them at the IS. We found previously that Nav1.6 is the predominant Nav channel isoform at IS of adult rodent retinal ganglion cells. However, here we report that Nav1.6 immunostaining is consistently reduced or absent in short regions of the IS proximal to the soma, although both ankyrin‐G and pan‐Nav antibodies stain this region. We show that this proximal IS subregion is a unique axonal microdomain, containing an accumulation of Nav1.1 channels that are spatially segregated from the Nav1.6 channels of the distal IS. Additionally, we find that axonal Kv1.2 potassium channels are present within the distal IS, but are also excluded from the Nav1.1‐enriched proximal IS microdomain. Because ankyrin‐G was prominent in both proximal and distal subcompartments of the IS, where it colocalized with either Nav1.1 or Nav1.6, respectively, mechanisms other than association with ankyrin‐G must mediate differential targeting of Nav channel subtypes to achieve the spatial precision observed within the IS. This precise arrangement of ion channels within the axon initial segment is likely an important determinant of the firing properties of ganglion cells and other mammalian neurons. J. Comp. Neurol. 500:339–352, 2007.

Response features of parvalbumin-expressing interneurons suggest precise roles for subtypes of inhibition in visual cortex.

Inhibitory interneurons in the cerebral cortex include a vast array of subtypes, varying in their molecular signatures, electrophysiological properties, and connectivity patterns. This diversity suggests that individual inhibitory classes have unique roles in cortical circuits; however, their characterization to date has been limited to broad classifications including many subtypes. We used the Cre/LoxP system, specifically labeling parvalbumin(PV)-expressing interneurons in visual cortex of PV-Cre mice with red fluorescent protein (RFP), followed by targeted loose-patch recordings and two-photon imaging of calcium responses in vivo to characterize the visual receptive field properties of these cells. Despite their relative molecular and morphological homogeneity, we find that PV+ neurons have a diversity of feature-specific visual responses that include sharp orientation and direction-selectivity, small receptive fields, and band-pass spatial frequency tuning. These results suggest that subsets of parvalbumin interneurons are components of specific cortical networks and that perisomatic inhibition contributes to the generation of precise response properties.

Molecular mechanisms of experience-dependent plasticity in visual cortex

A remarkable amount of our current knowledge of mechanisms underlying experience-dependent plasticity during cortical development comes from study of the mammalian visual cortex. Recent advances in high-resolution cellular imaging, combined with genetic manipulations in mice, novel fluorescent recombinant probes, and large-scale screens of gene expression, have revealed multiple molecular mechanisms that underlie structural and functional plasticity in visual cortex. We situate these mechanisms in the context of a new conceptual framework of feed-forward and feedback regulation for understanding how neurons of the visual cortex reorganize their connections in response to changes in sensory inputs. Such conceptual advances have important implications for understanding not only normal development but also pathological conditions that afflict the central nervous system.

Impaired Firing and Cell-Specific Compensation in Neurons Lacking Nav1.6 Sodium Channels

The ability of neurons to fire precise patterns of action potentials is critical for encoding inputs and efficiently driving target neurons. At the axon initial segment and nodes of Ranvier, where nerve impulses are generated and propagated, a high density of Nav1.2 sodium channels is developmentally replaced by Nav1.6 channels. In retinal ganglion cells (GCs), this isoform switch coincides with the developmental transition from single spikes to repetitive firing. Also, Nav1.6 channels are required for repetitive spiking in cerebellar Purkinje neurons. These previous observations suggest that the developmental appearance of Nav1.6 underlies the transition to repetitive spiking in GCs. To test this possibility, we recorded from GCs of med (Nav1.6-null) and wild-type mice during postnatal development. By postnatal day 18, when the switch to Nav1.6 at GC initial segments is normally complete, the maximal sustained and instantaneous firing rates were lower in med than in wild-type GCs, demonstrating that Nav1.6 channels are necessary to attain physiologically relevant firing frequencies in GCs. However, the firing impairment was milder than that reported previously in med Purkinje neurons, which prompted us to look for differences in compensatory sodium channel expression. Both Nav1.2 and Nav1.1 channels accumulated at initial segments and nodes of med GCs, sites normally occupied by Nav1.6. In med Purkinje cells, only Nav1.1 channels were found at initial segments, whereas in other brain regions, only Nav1.2 was detected at med initial segments and nodes. Thus, compensatory mechanisms in channel isoform distribution are cell specific, which likely results in different firing properties.

Gene expression patterns in visual cortex during the critical period: Synaptic stabilization and reversal by visual deprivation

The mapping of eye-specific, geniculocortical inputs to primary visual cortex (V1) is highly sensitive to the balance of correlated activity between the two eyes during a restricted postnatal critical period for ocular dominance plasticity. This critical period is likely to have amplified expression of genes and proteins that mediate synaptic plasticity. DNA microarray analysis of transcription in mouse V1 before, during, and after the critical period identified 31 genes that were up-regulated and 22 that were down-regulated during the critical period. The highest-ranked up-regulated gene, cardiac troponin C, codes for a neuronal calcium-binding protein that regulates actin binding and whose expression is activity-dependent and relatively selective for layer-4 star pyramidal neurons. The highest-ranked down-regulated gene, synCAM, also has actin-based function. Actin-binding function, G protein signaling, transcription, and myelination are prominently represented in the critical period transcriptome. Monocular deprivation during the critical period reverses the expression of nearly all critical period genes. The profile of regulated genes suggests that synaptic stability is a principle driver of critical period gene expression and that alteration in visual activity drives homeostatic restoration of stability.

Novel clustering of sodium channel Nav1.1 with ankyrin-G and neurofascin at discrete sites in the inner plexiform layer of the retina

Voltage-gated sodium channels cluster at sites of action potential generation and propagation by interacting with partner proteins such as neurofascin, an adhesion molecule in the L1 family, and ankyrin-G, a spectrin-binding protein required for sodium channel accumulation at axon initial segments. Here, we describe in the inner plexiform layer of the retina a novel site of high-density sodium channel clustering, marked by ankyrin-G and neurofascin. The sodium channel isoform at this site is Na(v)1.1, instead of the Na(v)1.6 channels more commonly found in association with the clustering machinery. During development, Na(v)1.2 channels first associate with ankyrin-G in the inner plexiform layer but are later replaced by Na(v)1.1, similar to the switch from Na(v)1.2 to Na(v)1.6 at nodes of Ranvier and initial segments. This represents the first instance of high-density clustering of Na(v)1.1 channels, which may contribute to synaptic interactions among retinal neurons in the inner plexiform layer.

Intrinsic patterning and experience-dependent mechanisms that generate eye-specific projections and binocular circuits in the visual pathway

A defining feature of the mammalian nervous system is its complex yet precise circuitry. The mechanisms which underlie the generation of neural connectivity are the topic of intense study in developmental neuroscience. The mammalian visual pathway demonstrates precise retinotopic organization in subcortical and cortical pathways, together with the alignment and matching of eye-specific projections, and sophisticated cortical circuitry that enables the extraction of features underlying vision. New approaches employing molecular-genetic analyses, transgenic mice, novel recombinant probes, and high-resolution imaging are contributing to rapid progress and a new synthesis in the field. These approaches are revealing the ways in which intrinsic patterning mechanisms act in concert with experience-dependent mechanisms to shape visual projections and circuits.

Expression of sodium channels Nav1.2 and Nav1.6 during postnatal development of the retina

During the second and third postnatal weeks, there is a developmental switch from sodium channel isoform Na(v)1.2 to isoform Na(v)1.6 at initial segments and nodes of Ranvier in rat retinal ganglion cells. We used quantitative, real-time PCR to determine if the developmental appearance of Na(v)1.6 channels is accompanied by an increase in steady-state level of Na(v)1.6 mRNA in the retina. Between postnatal day 2 (P2) and P10, Na(v)1.6 levels did not change, but between P10 and P19, there was an approximately three-fold increase in Na(v)1.6 transcript levels. This coincides with the appearance of Na(v)1.6 channels in the retina and optic nerve. The steady-state level of Na(v)1.2 mRNA also increased during this same period, which suggests that the rise in Na(v)1.6 may be part of a general increase in sodium channel transcripts at about the time of eye opening at P14. The results are consistent with a developmental increase in steady-state transcripts giving rise to a corresponding increase in sodium channel protein expression.

Influence of a Subtype of Inhibitory Interneuron on Stimulus-Specific Responses in Visual Cortex

Inhibition modulates receptive field properties and integrative responses of neurons in cortical circuits. The contribution of specific interneuron classes to cortical circuits and emergent responses is unknown. Here, we examined neuronal responses in primary visual cortex (V1) of adult Dlx1(-/-) mice, which have a selective reduction in cortical dendrite-targeting interneurons (DTIs) that express calretinin, neuropeptide Y, and somatostatin. The V1 neurons examined in Dlx1(-/-) mice have reduced orientation selectivity and altered firing rates, with elevated late responses, suggesting that local inhibition at dendrites has a specific role in modulating neuronal computations. We did not detect overt changes in the physiological properties of thalamic relay neurons and features of thalamocortical projections, such as retinotopic maps and eye-specific inputs, in the mutant mice, suggesting that the defects are cortical in origin. These experimental results are well explained by a computational model that integrates broad tuning from dendrite-targeting and narrower tuning from soma-targeting interneuron subclasses. Our findings suggest a key role for DTIs in the fine-tuning of stimulus-specific cortical responses.

STAT1 Regulates the Homeostatic Component of Visual Cortical Plasticity via an AMPA Receptor-Mediated Mechanism

Accumulating evidence points to a role for Janus kinase/signal transducers and activators of transcription (STAT) immune signaling in neuronal function; however, its role in experience-dependent plasticity is unknown. Here we show that one of its components, STAT1, negatively regulates the homeostatic component of ocular dominance plasticity in visual cortex. After brief monocular deprivation (MD), STAT1 knock-out (KO) mice show an accelerated increase of open-eye responses, to a level comparable with open-eye responses after a longer duration of MD in wild-type (WT) mice. Therefore, this component of plasticity is abnormally enhanced in KO mice. Conversely, increasing STAT1 signaling by IFNγ treatment in WT mice reduces the homeostatic component of plasticity by impairing open-eye responses. Enhanced plasticity in KO mice is accompanied by sustained surface levels of GluA1 AMPA receptors and increased amplitude and frequency of AMPA receptor-mediated mEPSCs, which resemble changes in WT mice after a longer duration of MD. These results demonstrate a unique role for STAT1 during visual cortical plasticity in vivo through a mechanism that includes AMPA receptors.