Aberrant hippocampal transmission and behavior in mice with a stargazin mutation linked to intellectual disability

Mutations linked to neurodevelopmental disorders, such as intellectual disability (ID), are frequently found in genes that encode for proteins of the excitatory synapse. Transmembrane AMPA receptor regulatory proteins (TARPs) are AMPA receptor auxiliary proteins that regulate crucial aspects of receptor function. Here, we investigate an ID-associated mutant form of the TARP family member stargazin. Molecular dynamics analyses showed that the stargazin V143L variant weakens the overall interface of the AMPAR:stargazin complex and hinders the stability of the complex. Knock-in mice for the V143L stargazin mutation manifest cognitive and social deficits and hippocampal synaptic transmission defects. In the hippocampus of stargazin V143L mice, CA1 neurons show impaired spine maturation in basal dendrites, and synaptic ultrastructural alterations. These data demonstrate a causal role for mutated stargazin in the pathogenesis of ID and highlight its role in the development and function of hippocampal synapses.

Gabra2 is a genetic modifier of Dravet syndrome in mice

Pathogenic variants in epilepsy genes result in a spectrum of clinical severity. One source of phenotypic heterogeneity is modifier genes that affect expressivity of a primary pathogenic variant. Mouse epilepsy models also display varying degrees of clinical severity on different genetic backgrounds. Mice with heterozygous deletion of Scn1a (Scn1a+/−) model Dravet syndrome, a severe epilepsy most often caused by SCN1A haploinsufficiency. Scn1a+/− mice recapitulate features of Dravet syndrome, including spontaneous seizures, sudden death, and cognitive/behavioral deficits. Scn1a+/− mice maintained on the 129S6/SvEvTac (129) strain have normal lifespan and no spontaneous seizures. In contrast, admixture with C57BL/6J (B6) results in epilepsy and premature lethality. We previously mapped Dravet Survival Modifier loci (Dsm1-Dsm5) responsible for strain-dependent differences in survival. Gabra2, encoding the GABAA α2 subunit, was nominated as a candidate modifier at Dsm1. Direct measurement of GABAA receptors found lower abundance of α2-containing receptors in hippocampal synapses of B6 mice relative to 129. We also identified a B6-specific single nucleotide deletion within Gabra2 that lowers mRNA and protein by nearly 50%. Repair of this deletion reestablished normal levels of Gabra2 expression. In this study, we used B6 mice with a repaired Gabra2 allele to evaluate Gabra2 as a genetic modifier of severity in Scn1a+/− mice. Gabra2 repair restored transcript and protein expression, increased abundance of α2-containing GABAA receptors in hippocampal synapses, and rescued epilepsy phenotypes of Scn1a+/− mice. These findings validate Gabra2 as a genetic modifier of Dravet syndrome, and support enhancing function of α2-containing GABAA receptors as treatment strategy for Dravet syndrome.

Sustained Hippocampal Synaptic Pathophysiology Following Single and Repeated Closed-Head Concussive Impacts

Traumatic brain injury (TBI), and related diseases such as chronic traumatic encephalopathy (CTE) and Alzheimer’s (AD), are of increasing concern in part due to enhanced awareness of their long-term neurological effects on memory and behavior. Repeated concussions, vs. single concussions, have been shown to result in worsened and sustained symptoms including impaired cognition and histopathology. To assess and compare the persistent effects of single or repeated concussive impacts on mediators of memory encoding such as synaptic transmission, plasticity, and cellular Ca2+ signaling, a closed-head controlled cortical impact (CCI) approach was used which closely replicates the mode of injury in clinical cases. Adult male rats received a sham procedure, a single impact, or three successive impacts at 48-hour intervals. After 30 days, hippocampal slices were prepared for electrophysiological recordings and 2-photon Ca2+ imaging, or fixed and immunostained for pathogenic phospho-tau species. In both concussion groups, hippocampal circuits showed hyper-excitable synaptic responsivity upon Schaffer collateral stimulation compared to sham animals, indicating sustained defects in hippocampal circuitry. This was not accompanied by sustained LTP deficits, but resting Ca2+ levels and voltage-gated Ca2+ signals were elevated in both concussion groups, while ryanodine receptor-evoked Ca2+ responses decreased with repeat concussions. Furthermore, pathogenic phospho-tau staining was progressively elevated in both concussion groups, with spreading beyond the hemisphere of injury, consistent with CTE. Thus, single and repeated concussions lead to a persistent upregulation of excitatory hippocampal synapses, possibly through changes in postsynaptic Ca2+ signaling/regulation, which may contribute to histopathology and detrimental long-term cognitive symptoms.

Regulation of hippocampal excitatory synapses by the Zdhhc5 palmitoyl acyl transferase

Palmitoylation is the most common post-translational lipid modification in the brain; however, the role of palmitoylation and palmitoylating enzymes in the nervous system remains elusive. One of these enzymes, Zdhhc5, has previously been shown to regulate synapse plasticity. Here, we report that Zdhhc5 is also essential for the formation of excitatory, but not inhibitory synapses both in vitro and in vivo. We demonstrate in vitro that this is dependent on Zdhhc5's enzymatic activity, its localization at the plasma membrane, and its C-terminal domain which has been shown to be truncated in a patient with schizophrenia. Loss of Zdhhc5 in mice results in a decrease in the density of excitatory hippocampal synapses accompanied by alterations in membrane capacitance and synaptic currents, consistent with an overall decrease in spine number and silent synapses. These findings reveal an important role for Zdhhc5 in the formation and/or maintenance of excitatory synapses.

Tacrolimus Decreases Cognitive Function by Impairing Hippocampal Synaptic Balance: A Possible Role of Klotho

The influence of long-term tacrolimus treatment on cognitive function remains to be elucidated. Using chronic tacrolimus neurotoxicity in mice, we evaluated the influence of tacrolimus on cognitive function, synaptic balance, its regulating protein (Klotho), and oxidative stress in the hippocampus. Compared to vehicle-treated mice, tacrolimus-treated mice showed significantly decreased hippocampal-dependent spatial learning and memory function. Furthermore, tacrolimus caused synaptic imbalance as demonstrated by decreased excitatory synapses and increased inhibitory synapses, and downregulated Klotho in a dose-dependent manner; its downregulation was localized to excitatory hippocampal synapses. Moreover, tacrolimus increased oxidative stress and was associated with the activation of the PI3K/AKT pathway in the hippocampus. The present results indicate that tacrolimus impairs cognitive function via synaptic imbalance, and that these processes are associated with Klotho downregulation at synapses through tacrolimus-induced oxidative stress in the hippocampus.