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Importance: The risk of mental disorders is consistently associated with variants in CACNA1C (L-type calcium channel Cav1.2) but it is not known why these channels are critical to cognition, and whether they affect the layer III pyramidal cells in the dorsolateral prefrontal cortex that are especially vulnerable in cognitive disorders., Objective: To examine the molecular mechanisms expressed in layer III pyramidal cells in primate dorsolateral prefrontal cortices., Design, Setting, and Participants: The design included transcriptomic analyses from human and macaque dorsolateral prefrontal cortex, and connectivity, protein expression, physiology, and cognitive behavior in macaques. The research was performed in academic laboratories at Yale, Harvard, Princeton, and the University of Pittsburgh. As dorsolateral prefrontal cortex only exists in primates, the work evaluated humans and macaques., Main Outcomes and Measures: Outcome measures included transcriptomic signatures of human and macaque pyramidal cells, protein expression and interactions in layer III macaque pyramidal cells using light and electron microscopy, changes in neuronal firing during spatial working memory, and working memory performance following pharmacological treatments., Results: Layer III pyramidal cells in dorsolateral prefrontal cortex coexpress a constellation of calcium-related proteins, delineated by CALB1 (calbindin), and high levels of CACNA1C (Cav1.2), GRIN2B (NMDA receptor GluN2B), and KCNN3 (SK3 potassium channel), concentrated in dendritic spines near the calcium-storing smooth endoplasmic reticulum. L-type calcium channels influenced neuronal firing needed for working memory, where either blockade or increased drive by β1-adrenoceptors, reduced neuronal firing by a mean (SD) 37.3% (5.5%) or 40% (6.3%), respectively, the latter via SK potassium channel opening. An L-type calcium channel blocker or β1-adrenoceptor antagonist protected working memory from stress., Conclusions and Relevance: The layer III pyramidal cells in the dorsolateral prefrontal cortex especially vulnerable in cognitive disorders differentially express calbindin and a constellation of calcium-related proteins including L-type calcium channels Cav1.2 (CACNA1C), GluN2B-NMDA receptors (GRIN2B), and SK3 potassium channels (KCNN3), which influence memory-related neuronal firing. The finding that either inadequate or excessive L-type calcium channel activation reduced neuronal firing explains why either loss- or gain-of-function variants in CACNA1C were associated with increased risk of cognitive disorders. The selective expression of calbindin in these pyramidal cells highlights the importance of regulatory mechanisms in neurons with high calcium signaling, consistent with Alzheimer tau pathology emerging when calbindin is lost with age and/or inflammation.

Most mental disorders involve dysfunction of the dorsolateral prefrontal cortex (dlPFC), a recently evolved brain region that subserves working memory, abstraction, and the thoughtful regulation of attention, action, and emotion. For example, schizophrenia, depression, long COVID, and Alzheimer's disease are all associated with dlPFC dysfunction, with neuropathology often being focused in layer III. The dlPFC has extensive top-down projections, e.g., to the posterior association cortices to regulate attention and to the subgenual cingulate cortex via the rostral and medial PFC to regulate emotional responses. However, the dlPFC is particularly dependent on arousal state and is very vulnerable to stress and inflammation, which are etiological and/or exacerbating factors for most mental disorders. The cellular mechanisms by which stress and inflammation impact the dlPFC are a topic of current research and are summarized in this review. For example, the layer III dlPFC circuits that generate working memory-related neuronal firing have unusual neurotransmission, depending on NMDA receptor and nicotinic α7 receptor actions that are blocked under inflammatory conditions by kynurenic acid. These circuits also have unusual neuromodulation, with the molecular machinery to magnify calcium signaling in spines needed to support persistent firing, which must be tightly regulated to prevent toxic calcium actions. Stress rapidly weakens layer III connectivity by driving feedforward calcium-cAMP (cyclic adenosine monophosphate) opening of potassium channels on spines. This is regulated by postsynaptic noradrenergic α 2A adrenergic receptor and mGluR3 (metabotropic glutamate receptor 3) signaling but dysregulated by inflammation and/or chronic stress exposure, which contribute to spine loss. Treatments that strengthen the dlPFC via pharmacological (the α 2A adrenergic receptor agonist, guanfacine) or repetitive transcranial magnetic stimulation manipulation provide a rational basis for therapy., (Copyright © 2024 Society of Biological Psychiatry. Published by Elsevier Inc. All rights reserved.)

The human GlyT1 glycine transporter requires chloride for its function. However, the mechanism by which Cl - exerts its influence is unknown. To examine the role that Cl - plays in the transport cycle, we measured the effect of Cl - on both glycine binding and conformational changes. The ability of glycine to displace the high-affinity radioligand [ 3 H]CHIBA-3007 required Na + and was potentiated over 1,000-fold by Cl - We generated GlyT1b mutants containing reactive cysteine residues in either the extracellular or cytoplasmic permeation pathways and measured changes in the reactivity of those cysteine residues as indicators of conformational changes in response to ions and substrate. Na + increased accessibility in the extracellular pathway and decreased it in the cytoplasmic pathway, consistent with stabilizing an outward-open conformation as observed in other members of this transporter family. In the presence of Na + , both glycine and Cl - independently shifted the conformation of GlyT1b toward an outward-closed conformation. Together, Na + , glycine, and Cl - stabilized an inward-open conformation of GlyT1b. We then examined whether Cl - acts by interacting with a conserved glutamine to allow formation of an ion pair that stabilizes the closed state of the extracellular pathway. Molecular dynamics simulations of a GlyT1 homolog indicated that this ion pair is formed more frequently as that pathway closes. Mutation of the glutamine blocked the effect of Cl - , and substituting it with glutamate or lysine resulted in outward- or inward-facing transporter conformations, respectively. These results provide an unexpected insight into the role of Cl - in this family of transporters., Competing Interests: The authors declare no competing interest.