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The transcriptional coactivator, PGC-1α (peroxisome proliferator-activated receptor γ coactivator 1α), plays a key role in coordinating energy requirement within cells. Its importance is reflected in the growing number of psychiatric and neurological conditions that have been associated with reduced PGC-1α levels. In cortical networks, PGC-1α is required for the induction of parvalbumin (PV) expression in interneurons, and PGC-1α deficiency affects synchronous GABAergic release. It is unknown, however, how this affects cortical excitability. We show here that knocking down PGC-1α specifically in the PV-expressing cells (PGC-1αPV−/−) blocks the activity-dependent regulation of the synaptic proteins, SYT2 and CPLX1. More surprisingly, this cell class-specific knockout of PGC-1α appears to have a novel antiepileptic effect, as assayed in brain slices bathed in 0 Mg2+ media. The rate of occurrence of preictal discharges developed approximately equivalently in wild-type and PGC-1αPV−/− brain slices, but the intensity of these discharges was lower in PGC-1αPV−/− slices, as evident from the reduced power in the γ range and reduced firing rates in both PV interneurons and pyramidal cells during these discharges. Reflecting this reduced intensity in the preictal discharges, the PGC-1αPV−/− brain slices experienced many more discharges before transitioning into a seizure-like event. Consequently, there was a large increase in the latency to the first seizure-like event in brain slices lacking PGC-1α in PV interneurons. We conclude that knocking down PGC-1α limits the range of PV interneuron firing and this slows the pathophysiological escalation during ictogenesis. NEW & NOTEWORTHY Parvalbumin expressing interneurons are considered to play an important role in regulating cortical activity. We were surprised, therefore, to find that knocking down the transcriptional coactivator, PGC-1α, specifically in this class of interneurons appears to slow ictogenesis. This anti-ictogenic effect is associated with reduced activity in preictal discharges, but with a far longer period of these discharges before the first seizure-like events finally start. Thus, PGC-1α knockdown may promote schizophrenia while reducing epileptic tendencies.

The transcriptional coactivator, PGC-1α (peroxisome proliferator activated receptor gamma coactivator 1α), plays a key role coordinating energy requirement within cells. Its importance is reflected in the growing number of psychiatric and neurological conditions that have been associated with reduced PGC-1α levels. In cortical networks, PGC-1α is required for the induction of parvalbumin (PV) expression in interneurons, and PGC-1α deficiency affects synchronous GABAergic release. It is unknown, however, how this affects cortical excitability. We show here that knocking down PGC-1α specifically in the PV-expressing cells (PGC-1αPV-/-), blocks the activity-dependent regulation of the synaptic proteins, SYT2 and CPLX1. More surprisingly, this cell-class specific knock-out of PGC-1α appears to have a novel anti-epileptic effect, as assayed in brain slices bathed in 0 Mg2+ media. The rate of pre-ictal discharges developed approximately equivalently in wild-type and PGC-1αPV-/- brain slices, but the intensity of these discharges was lower in PGC-1αPV-/- slices, as evident from the reduced power in the gamma range and reduced firing rates in both PV interneurons and pyramidal cells during these discharges. Reflecting this reduced intensity in the pre-ictal discharges, the PGC-1αPV-/- brain slices experienced many more discharges before transitioning into a seizure-like event. Consequently, there was a large increase in the latency to the first seizure-like event in brain slices lacking PGC-1α in PV interneurons. We conclude that knocking down PGC-1α limits the range of PV interneuron firing, and this slows the pathophysiological escalation during ictogenesis.

We report the case of a patient with a clinical phenotype characterized by distal lower limb weakness and pes cavus. The electrophysiological study showed slightly reduced or normal amplitude of motor potentials, a decremental response to repetitive nerve stimulation and post-exercise facilitation. Muscle biopsy showed only mild neurogenic features. Genetic analysis included a clinical exome sequencing, followed by Sanger analysis. Three-dimensional (3D) models were generated with a SwissModel (https://swissmodel.expasy.org/) to explain the clinical observations and reinforce the pathogenic nature of the genetic variant identified. Genetic analysis demonstrated a new de novo heterozygous in frame deletion of the SYT2 gene (NM_177402.4: c.1082_1096del), confirmed by Sanger sequencing, which removes five aminoacids in the C2B domain of synaptotagmin-2 protein, that cause a profound effect on the structure and function of this synaptic vesicle protein. We identified a de novo genetic variant in the SYT2 gene, further supporting its association with a highly stereotyped clinical and electrophysiological phenotype. Our case showed electrophysiological features consistent with a presynaptic dysfunction in the neuromuscular junction with normal post-exercise amplitudes, not supporting the presence of predominant axonal damage. Although the analysis of SYT2 gene should be included in genetic analysis of patients presenting with this clinical phenotype that mimics motor neuropathy, clinicians have to consider the study of neuromuscular transmission to early identify this potentially treatable condition.

Objective: To report the first de novo missense mutation in the SYT2 gene causing distal hereditary motor neuropathy. Methods: Genetic testing was carried out, including clinical exome sequencing for the proband and Sanger sequencing for the proband and his parents. We described the clinical and electrophysiological features found in the patient. Results: We reported a proband with a new de novo missense mutation, c.917C>T (p.Ser306Leu), in the C2B domain of SYT2. The clinical presentation was similar to that of phenotypes described in previous studies. A notable feature in our study was normal electrophysiological testing results of the patient. Conclusions: In this study we reinforced the association between SYT2 mutations and distal hereditary motor neuropathy. We also described the clinical presentation of the patient carrying this pathogenic variant and provided unusual results of electrophysiological testing. The results showed that a diagnosis of SYT2-associated neuropathy should be based on the similarity of clinical manifestations, rather than the results of electrophysiological testing.

Regulated secretion is controlled by Ca2+sensors with different affinities and subcellular distributions. Inactivation ofSyt1(synaptotagmin-1), the main Ca2+sensor for synchronous neurotransmission in many neurons, enhances asynchronous and spontaneous release rates, suggesting that Syt1 inhibits other sensors with higher Ca2+affinities and/or lower cooperativities. Such sensors could include Doc2a and Doc2b, which have been implicated in spontaneous and asynchronous neurotransmitter release and compete with Syt1 for binding SNARE complexes. Here, we tested this hypothesis using triple-knock-out mice. Inactivation of Doc2a and Doc2b in Syt1-deficient neurons did not reduce the high spontaneous release rate. Overexpression of Doc2b variants in triple-knock-out neurons reduced spontaneous release but did not rescue synchronous release. A chimeric construct in which the C2AB domain of Syt1 was substituted by that of Doc2b did not support synchronous release either. Conversely, the soluble C2AB domain of Syt1 did not affect spontaneous release. We conclude that the high spontaneous release rate in synaptotagmin-deficient neurons does not involve the binding of Doc2 proteins to Syt1 binding sites in the SNARE complex. Instead, our results suggest that the C2AB domains of Syt1 and Doc2b specifically support synchronous and spontaneous release by separate mechanisms. (Both male and female neurons were studied without sex determination.)SIGNIFICANCE STATEMENTNeurotransmission in the brain is regulated by presynaptic Ca2+concentrations. Multiple Ca2+sensor proteins contribute to synchronous (Syt1, Syt2), asynchronous (Syt7), and spontaneous (Doc2a/Doc2b) phases of neurotransmitter release. Genetic ablation of synchronous release was previously shown to affect other release phases, suggesting that multiple sensors may compete for similar release sites, together encoding stimulus–secretion coupling over a large range of synaptic Ca2+concentrations. Here, we investigated the extent of functional overlap between Syt1, Doc2a, and Doc2b by reintroducing wild-type and mutant proteins in triple-knock-out neurons, and conclude that the sensors are highly specialized for different phases of release.

Contains fulltext : 229372.pdf (Publisher’s version ) (Closed access) Synaptotagmins are integral synaptic vesicle membrane proteins that function as calcium sensors and regulate neurotransmitter release at the presynaptic nerve terminal. Synaptotagmin-2 (SYT2), is the major isoform expressed at the neuromuscular junction. Recently, dominant missense variants in SYT2 have been reported as a rare cause of distal motor neuropathy and myasthenic syndrome, manifesting with stable or slowly progressive distal weakness of variable severity along with presynaptic NMJ impairment. These variants are thought to have a dominant-negative effect on synaptic vesicle exocytosis, although the precise pathomechanism remains to be elucidated. Here we report seven patients of five families, with biallelic loss of function variants in SYT2, clinically manifesting with a remarkably consistent phenotype of severe congenital onset hypotonia and weakness, with variable degrees of respiratory involvement. Electrodiagnostic findings were consistent with a presynaptic congenital myasthenic syndrome (CMS) in some. Treatment with an acetylcholinesterase inhibitor pursued in three patients showed clinical improvement with increased strength and function. This series further establishes SYT2 as a CMS-disease gene and expands its clinical and genetic spectrum to include recessive loss-of-function variants, manifesting as a severe congenital onset presynaptic CMS with potential treatment implications.

The objective: check the assumption that stem cells with modified adhesive properties (SC-m-SYT1/SYT2) should be more pronounced than native SCs, restore the phosphatase and contractile activity of cavernous tissue damaged by hyperglycemia. Materials and methods. A comparative study of the effect on the contractile and phosphate activity of the cavernous tissue of four intra-cavernous injections (ICI) (1106 cells once a week) of native stem cells (SC) and cells, with SYT1/SYT2 molecules, integrated into the cell membrane (SC-m-SYT1/SYT2), was performed in two groups (n=10; 1:1) of sexually mature Wistar rats weighing 160-180 g with induced hyperglycemia (>8 mmol/L). Results. It was found that, at the end of the ICI in the group of animals receiving SC-m-SYT1/SYT2, the median amplitude of contractions was 1.6 times greater (р

The human frontal cortex is unusually large compared with many other species. The expansion of the human frontal cortex is accompanied by both connectivity and transcriptional changes. Yet, the developmental origins generating variation in frontal cortex circuitry across species remain unresolved. Nineteen genes, which encode filaments, synapse, and voltage-gated channels (e.g., NEFH, SYT2, VAMP1) are especially enriched in the supragranular layers of the cerebral cortex in humans relative to mice. The increased expression of these genes suggests enhanced cortico-cortical projections emerging from layer III in humans. We confirm that the expression of these supragranular-enriched genes is preferentially expressed in frontal cortex layer III in humans relative to mice. We demonstrate a concomitant expansion in cortico-cortical pathways projecting within the frontal cortex white matter in humans with diffusion MR tractography. To identify developmental sources of such variation, we compare frontal cortical white matter growth and developmental trajectories of transcriptional profiles of supragranular-enriched genes in humans and mice. We also use temporal changes in gene expression during postnatal development to control for variation in developmental schedules across species. The growth of the frontal cortex white matter and transcriptional profiles of supragranular genes are both protracted in humans relative to the timing of other transformations. These findings demonstrate that an expansion of projections emerging from the human frontal cortex is achieved by extending the duration of cortical circuitry development. Integrating RNA sequencing with neuroimaging level phenotypes is an effective strategy to assess deviations in developmental programs leading to variation in connections across species.

Parvalbumin-positive (PV+) interneurons play a pivotal role in orchestrating windows of experience-dependent brain plasticity during development. Critical period closure is marked by the condensation of a perineuronal net (PNN) tightly enwrapping subsets of PV+ neurons, both acting as a molecular brake on plasticity and maintaining mature PV+ cell signaling. As much of the molecular organization of PNNs exists at length scales near or below the diffraction limit of light microscopy, we developed a superresolution imaging and analysis platform to visualize the structural organization of PNNs and the synaptic inputs perforating them in primary visual cortex. We identified a structural trajectory of PNN maturation featuring a range of net structures, which was accompanied by an increase in Synaptotagmin-2 (Syt2) signals on PV+ cells suggestive of increased inhibitory input between PV+ neurons. The same structural trajectory was followed by PNNs both during normal development and under conditions of critical period delay by total sensory deprivation or critical period acceleration by deletion of MeCP2, the causative gene for Rett syndrome, despite shifted maturation levels under these perturbations. Notably, superresolution imaging further revealed a decrease in Syt2 signals alongside an increase in vesicular glutamate transporter-2 signals on PV+ cells in MeCP2-deficient animals, suggesting weaker recurrent inhibitory input between PV+ neurons and stronger thalamocortical excitatory inputs onto PV+ cells. These results imply a latent imbalanced circuit signature that might promote cortical silencing in Rett syndrome before the functional regression of vision.

One in 26 people develop epilepsy and in these temporal lobe epilepsy (TLE) is common. Many patients display a pattern of neuron loss called hippocampal sclerosis. Seizures usually start in the hippocampus but underlying mechanisms remain unclear. One possibility is insufficient inhibition of dentate granule cells. Normally parvalbumin-immunoreactive (PV) interneurons strongly inhibit granule cells. Humans with TLE display loss of PV interneurons in the dentate gyrus but questions persist. To address this, we evaluated PV interneuron and bouton numbers in California sea lions (Zalophus californianus) that naturally develop TLE after exposure to domoic acid, a neurotoxin that enters the marine food chain during harmful algal blooms. Sclerotic hippocampi were identified by the loss of Nissl-stained hilar neurons. Stereological methods were used to estimate the number of granule cells and PV interneurons per dentate gyrus. Sclerotic hippocampi contained fewer granule cells, fewer PV interneurons, and fewer PV synaptic boutons, and the ratio of granule cells to PV interneurons was higher than in controls. To test whether fewer boutons was attributable to loss versus reduced immunoreactivity, expression of synaptotagmin-2 (syt2) was evaluated. Syt2 is also expressed in boutons of PV interneurons. Sclerotic hippocampi displayed proportional losses of syt2-immunoreactive boutons, PV boutons, and granule cells. There was no significant difference in the average numbers of PV- or syt2-positive boutons per granule cell between control and sclerotic hippocampi. These findings do not address functionality of surviving synapses but suggest reduced granule cell inhibition in TLE is not attributable to anatomical loss of PV boutons.

The voltage-gated sodium channel NaV1.7 plays a critical role in pain pathways. Besides action potential propagation, NaV1.7 regulates neurotransmitter release, integrates depolarizing inputs over long periods and regulates transcription. In order to better understand these functions, we generated an epitope-tagged NaV1.7 mouse that showed normal pain behavior. Analysis of NaV1.7 complexes affinity-purified under native conditions by mass spectrometry revealed 267 NaV1.7 associated proteins including known interactors, such as the sodium channel β3 subunit (Scn3b) and collapsin response mediator protein (Crmp2), and novel interactors. Selected novel NaV1.7 protein interactors membrane-trafficking protein synapototagmin-2 (Syt2), G protein-regulated inducer of neurite outgrowth 1 (Gprin1), L-type amino acid transporter 1 (Lat1) and transmembrane P24 trafficking protein 10 (Tmed10) together with Scn3b and Crmp2 were validated using co-immunoprecipitation and functional assays. The information provided with this physiologically normal epitope-tagged mouse should provide useful insights into the pain mechanisms associated with NaV1.7 channel function.

Accumulating evidence strongly implicates the transcriptional coactivator peroxisome proliferator-activated receptor γ coactivator 1α (PGC-1α) in the pathophysiology of multiple neurological disorders, but the downstream gene targets of PGC-1α in the brain have remained enigmatic. Previous data demonstrate that PGC-1α is primarily concentrated in inhibitory neurons and that PGC-1α is required for the expression of the interneuron-specific Ca2+-binding protein parvalbumin (PV) throughout the cortex. To identify other possible transcriptional targets of PGC-1α in neural tissue, we conducted a microarray on neuroblastoma cells overexpressing PGC-1α, mined results for genes with physiological relevance to interneurons, and measured cortical gene and protein expression of these genes in mice with underexpression and overexpression of PGC-1α. We observed bidirectional regulation of novel PGC-1α-dependent transcripts spanning synaptic [synaptotagmin 2 (Syt2) and complexin 1 (Cplx1)], structural [neurofilament heavy chain (Nefh)], and metabolic [neutral cholesterol ester hydrolase 1 (Nceh1), adenylate kinase 1 (Ak1), inositol polyphosphate 5-phosphatase J (Inpp5j), ATP synthase mitochondrial F1 complex O subunit (Atp5o), phytanol-CoA-2hydroxylase (Phyh), and ATP synthase mitrochondrial F1 complex α subunit 1 (Atp5a1)] functions. The neuron-specific genes Syt2, Cplx1, and Nefh were developmentally upregulated in an expression pattern consistent with that of PGC-1α and were expressed in cortical interneurons. Conditional deletion of PGC-1α in PV-positive neurons significantly decreased cortical transcript expression of these genes, promoted asynchronous GABA release, and impaired long-term memory. Collectively, these data demonstrate that PGC-1α is required for normal PV-positive interneuron function and that loss of PGC-1α in this interneuron subpopulation could contribute to cortical dysfunction in disease states.

The voltage‐gated sodium channel Na V 1.7 plays a critical role in pain pathways. We generated an epitope‐tagged Na V 1.7 mouse that showed normal pain behaviours to identify channel‐interacting proteins. Analysis of Na V 1.7 complexes affinity‐purified under native conditions by mass spectrometry revealed 267 proteins associated with Nav1.7 in vivo . The sodium channel β3 (Scn3b), rather than the β1 subunit, complexes with Nav1.7, and we demonstrate an interaction between collapsing‐response mediator protein (Crmp2) and Nav1.7, through which the analgesic drug lacosamide regulates Nav1.7 current density. Novel Na V 1.7 protein interactors including membrane‐trafficking protein synaptotagmin‐2 (Syt2), L‐type amino acid transporter 1 (Lat1) and transmembrane P24‐trafficking protein 10 (Tmed10) together with Scn3b and Crmp2 were validated by co‐immunoprecipitation (Co‐IP) from sensory neuron extract. Nav1.7, known to regulate opioid receptor efficacy, interacts with the G protein‐regulated inducer of neurite outgrowth (Gprin1), an opioid receptor‐binding protein, demonstrating a physical and functional link between Nav1.7 and opioid signalling. Further information on physiological interactions provided with this normal epitope‐tagged mouse should provide useful insights into the many functions now associated with the Na V 1.7 channel.

Synaptotagmins are integral synaptic vesicle membrane proteins that function as calcium sensors and regulate neurotransmitter release at the presynaptic nerve terminal. Synaptotagmin-2 (SYT2), is the major isoform expressed at the neuromuscular junction. Recently, dominant missense variants in SYT2 have been reported as a rare cause of distal motor neuropathy and myasthenic syndrome, manifesting with stable or slowly progressive distal weakness of variable severity along with presynaptic NMJ impairment. These variants are thought to have a dominant-negative effect on synaptic vesicle exocytosis, although the precise pathomechanism remains to be elucidated. Here we report seven patients of five families, with biallelic loss of function variants in SYT2, clinically manifesting with a remarkably consistent phenotype of severe congenital onset hypotonia and weakness, with variable degrees of respiratory involvement. Electrodiagnostic findings were consistent with a presynaptic congenital myasthenic syndrome (CMS) in some. Treatment with an acetylcholinesterase inhibitor pursued in three patients showed clinical improvement with increased strength and function. This series further establishes SYT2 as a CMS-disease gene and expands its clinical and genetic spectrum to include recessive loss-of-function variants, manifesting as a severe congenital onset presynaptic CMS with potential treatment implications.

Various Synaptotagmin (Syt) isoform genes are found in mammals, but it is unknown whether Syts can function redundantly in a given nerve terminal, or whether isoforms can be switched during the development of a nerve terminal. Here, we investigated the possibility of a developmental Syt isoform switch using the calyx of Held as a model synapse. At mature calyx synapses, fast Ca(2+)-driven transmitter release depended entirely on Syt2, but the release phenotype of Syt2 knockout (KO) mice was weaker at immature calyces, and absent at pre-calyceal synapses early postnatally. Instead, conditional genetic inactivation shows that Syt1 mediates fast release at pre-calyceal synapses, as well as a fast release component resistant to Syt2 deletion in immature calyces. This demonstrates a developmental Syt1-Syt2 isoform switch at an identified synapse, a mechanism that could fine-tune the speed, reliability, and plasticity of transmitter release at fast releasing CNS synapses.