Tissue‐ and cell‐specific expression of a splice variant in the II‐III cytoplasmic loop of Cacna1b

Abstract

Presynaptic Ca V 2.2 (N‐type) channels are fundamental for transmitter release across the nervous system. The gene encoding Ca V 2.2 channels, Cacna1b , contains alternatively spliced exons that result in functionally distinct splice variants (e18a, e24a, e31a, and 37a/37b). Alternative splicing of the cassette exon 18a generates two mRNA transcripts (+e18a‐ Cacna1b and ∆e18a‐ Cacna1b ). In this study, using novel mouse genetic models and in situ hybridization (BaseScope™), we confirmed that +e18a‐ Cacna1b splice variants are expressed in monoaminergic regions of the midbrain. We expanded these studies and identified +e18a‐ Cacna1b mRNA in deep cerebellar cells and spinal cord motor neurons. Furthermore, we determined that +e18a ‐Cacna1b is enriched in cholecystokinin‐expressing interneurons. Our results provide key information to understand cell‐specific functions of Ca V 2.2 channels.

N-type calcium channels control GABAergic transmission in brain areas related to fear and anxiety

“…CCK + INs were labeled for histochemistry and electrophysiology using an intersectional labeling approach. Our results are similar to previous studies before by our group and others [27,41,45,48,49]. We found that IPSCs recorded on BLA small CCK + INs co-express VIP, but their function is not known [37,51].…”

“…The identity of these cells was previously confirmed using fluorescence activated cell sorting coupled to qRT-PCR. Cells expressing tdT also expressed GABAergic markers such as glutamate decarboxylase-2 mRNA and the cannabinoid receptor 1 mRNA; both are hallmarks of CCK + INs [27].…”

“…Mice were genotyped using toe biopsy as done previously [27]. Primer sequences and band size are located in Table 1.…”

“…Additional evidence implicating N-type calcium channels in anxiety arises from behavioral analysis of novel mouse genetic models, behavioral studies with brain infusions of w-conotoxin GVIA (w-ctx GVIA), and clinical studies in humans. Mice with restricted splice choice in the gene that encodes for the a1-pore forming subunit of CaV2.2, Cacna1b (lack 37a-Cacna1b splice variant), show alterations in exploratory behavior in basal conditions and under mild stress [27]. Infusions of wctx GVIA in the medial prefrontal cortex (mPFC) impairs recall of extinguished memories in fear conditioning, and ventricular application of w-ctx GVIA results in reduced exploratory behavior in the elevated plus maze and light-dark box [28,29].…”

“…In these mice, Cre-Lox and Flpe-FRT recombination removed the two STOP cassettes, resulting in tdT expression in CCK + INs. Successful labeling of CCK + INs was validated in a previous work from our lab[27].To perform electrophysiological studies of synapses between CCK + INs and PNs, we used optogenetics. CCK + INs were labeled with red-shifted channelrhodopsin (ReaChR) fused to m-Citrine.…”