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Voltage-gated calcium channels are important for thalamocortical (TC) oscillations related to spike-wave discharges (SWDs) during absence seizures. The role of Ca V 2.3 R-type channels expressed in the thalamic intralaminar complex in SWDs, however, is not well studied. We investigated pharmacologically induced SWDs from the central medial thalamus (CMT) and somatosensory cortex in a Ca V 2.3 knockout (KO) mouse model using local field potential (LFP), and electroencephalographic (EEG) recordings. The duration of cumulative SWDs was significantly decreased in Ca V 2.3 KO mice compared with wild-type (WT) mice. A characteristic increase in the delta and theta waves was observed in both the CMT and somatosensory cortex during SWDs with delta (1-4 Hz) band TC synchronization increasing only in WT animals. Specifically, in the KO mice, LFPs recorded from the CMT showed no significant changes in the delta band and a significant decrease in the theta (4-8 Hz) band, and cortical EEG recordings showed a significant increase in the delta band, but no changes in the theta band. The baseline TC phase synchronization in the delta band was also more pronounced in the Ca V 2.3 KO mice than in WT mice. These findings suggest that R-type calcium channels in the CMT play a crucial role in sustaining and promoting the oscillatory activity of the TC network during absence seizures., Competing Interests: Declarations. Competing interests: The authors declare no competing interests. Ethical statement: The experimental procedures involving animals in this study were conducted under a protocol approved by the Institutional Animal Care and Use Committee of the University of Colorado Anschutz Medical Campus, Aurora, CO, USA. The treatment of animals adhered to guidelines set forth in the NIH Guide for the Care and Use of Laboratory Animals., (© 2025. The Author(s).)

The Ca V 3.2 isoform of T-type voltage-gated calcium channels plays a crucial role in regulating the excitability of nociceptive neurons; the endogenous molecules that modulate its activity, however, remain poorly understood. Here, we used serum proteomics and patch-clamp physiology to discover a novel peptide albumin (1-26) that facilitates channel gating by chelating trace metals that tonically inhibit Ca V 3.2 via H191 residue. Importantly, serum also potently modulated T-currents in human and rodent dorsal root ganglion (DRG) neurons. In vivo pain studies revealed that injections of serum and albumin (1-26) peptide resulted in robust mechanical and heat hypersensitivity. This hypersensitivity was abolished with a T-channel inhibitor, in Ca V 3.2 null mice and in Ca V 3.2 H191Q knock-in mice. The discovery of endogenous chelators of trace metals in the serum deepens our understanding of the role of Ca V 3.2 channels in neuronal hyperexcitability and may facilitate the design of novel analgesics with unique mechanisms of action.

We recently reported that a neurosteroid analogue with T-channel-blocking properties (3β,5β,17β)-3-hydroxyandrostane-17-carbonitrile (3β-OH), induced hypnosis in rat pups without triggering neuronal apoptosis. Furthermore, we found that the inhibition of the Ca V 3.1 isoform of T-channels contributes to the hypnotic properties of 3β-OH in adult mice. However, the specific mechanisms underlying the role of other subtypes of voltage-gated calcium channels in thalamocortical excitability and oscillations in vivo during 3β-OH-induced hypnosis are largely unknown. Here, we used patch-clamp recordings from acute brain slices, in vivo electroencephalogram (EEG) recordings, and mouse genetics with wild-type (WT) and Ca V 2.3 knock-out (KO) mice to further investigate the molecular mechanisms of neurosteroid-induced hypnosis. Our voltage-clamp recordings showed that 3β-OH inhibited recombinant Ca V 2.3 currents. In subsequent current-clamp recordings in thalamic slices ex vivo , we found that selective Ca V 2.3 channel blocker (SNX-482) inhibited stimulated tonic firing and increased the threshold for rebound burst firing in WT animals. Additionally, in thalamic slices we found that 3β-OH inhibited spike-firing more profoundly in WT than in mutant mice. Furthermore, 3β-OH reduced bursting frequencies in WT but not mutant animals. In ensuing in vivo experiments, we found that intra-peritoneal injections of 3β-OH were less effective in inducing LORR in the mutant mice than in the WT mice, with expected sex differences. Furthermore, the reduction in total α, β, and low γ EEG power was more profound in WT than in Ca V 2.3 KO females over time, while at 60 min after injections of 3β-OH, the increase in relative β power was higher in mutant females. In addition, 3β-OH depressed EEG power more strongly in the male WT than in the mutant mice and significantly increased the relative δ power oscillations in WT male mice in comparison to the mutant male animals. Our results demonstrate for the first time the importance of the Ca V 2.3 subtype of voltage-gated calcium channels in thalamocortical excitability and the oscillations that underlie neurosteroid-induced hypnosis., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Timic Stamenic, Manzella, Maksimovic, Krishnan, Covey, Jevtovic-Todorovic and Todorovic.)

General anesthetics mainly act by modulating synaptic inhibition on the one hand (the potentiation of GABA transmission) or synaptic excitation on the other (the inhibition of NMDA receptors), but they can also have effects on numerous other proteins, receptors, and channels. The effects of general anesthetics on ion channels have been the subject of research since the publication of reports of direct actions of these drugs on ion channel proteins. In particular, there is considerable interest in T-type voltage-gated calcium channels that are abundantly expressed in the thalamus, where they control patterns of cellular excitability and thalamocortical oscillations during awake and sleep states. Here, we summarized and discussed our recent studies focused on the Ca V 3.1 isoform of T-channels in the nonspecific thalamus (intralaminar and midline nuclei), which acts as a key hub through which natural sleep and general anesthesia are initiated. We used mouse genetics and in vivo and ex vivo electrophysiology to study the role of thalamic T-channels in hypnosis induced by a standard general anesthetic, isoflurane, as well as novel neuroactive steroids. From the results of this study, we conclude that Ca V 3.1 channels contribute to thalamocortical oscillations during anesthetic-induced hypnosis, particularly the slow-frequency range of δ oscillations (0.5-4 Hz), by generating "window current" that contributes to the resting membrane potential. We posit that the role of the thalamic Ca V 3.1 isoform of T-channels in the effects of various classes of general anesthetics warrants consideration.

Background: Many neuroactive steroids induce sedation/hypnosis by potentiating γ-aminobutyric acid (GABA A ) currents. However, we previously demonstrated that an endogenous neuroactive steroid epipregnanolone [(3β,5β)-3-hydroxypregnan-20-one] (EpiP) exerts potent peripheral analgesia and blocks T-type calcium currents while sparing GABA A currents in rat sensory neurons. This study seeks to investigate the behavioral effects elicited by systemic administration of EpiP and to characterize its use as an adjuvant agent to commonly used general anesthetics (GAs)., Methods: Here, we utilized electroencephalographic (EEG) recordings to characterize thalamocortical oscillations, as well as behavioral assessment and mouse genetics with wild-type (WT) and different knockout (KO) models of T-channel isoforms to investigate potential sedative/hypnotic and immobilizing properties of EpiP., Results: Consistent with increased oscillations in slower EEG frequencies, EpiP induced an hypnotic state in WT mice when injected alone intra-peritoneally (i.p.) and effectively facilitated anesthetic effects of isoflurane (ISO) and sevoflurane (SEVO). The Ca V 3.1 (Cacna1g) KO mice demonstrated decreased sensitivity to EpiP-induced hypnosis when compared to WT mice, whereas no significant difference was noted between Ca V 3.2 (Cacna1h), Ca V 3.3 (Cacna1i) and WT mice. Finally, when compared to WT mice, onset of EpiP-induced hypnosis was delayed in Ca V 3.2 KO mice but not in Ca V 3.1 and Ca V 3.3 KO mice., Conclusion: We posit that EpiP may have an important role as novel hypnotic and/or adjuvant to volatile anesthetic agents. We speculate that distinct hypnotic effects of EpiP across all three T-channel isoforms is due to their differential expression in thalamocortical circuitry., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Background: The mechanisms underlying the role of T-type calcium channels (T-channels) in thalamocortical excitability and oscillations in vivo during neurosteroid-induced hypnosis are largely unknown., Methods: We used patch-clamp electrophysiological recordings from acute brain slices ex vivo, recordings of local field potentials (LFPs) from the central medial thalamic nucleus in vivo, and wild-type (WT) and Ca v 3.1 knock-out mice to investigate the molecular mechanisms of hypnosis induced by the neurosteroid analogue (3β,5β,17β)-3-hydroxyandrostane-17-carbonitrile (3β-OH)., Results: Patch-clamp recordings showed that 3β-OH inhibited isolated T-currents but had no effect on phasic or tonic γ-aminobutyric acid A currents. Also in acute brain slices, 3β-OH inhibited the spike firing mode more profoundly in WT than in Ca v 3.1 knockout mice. Furthermore, 3β-OH significantly hyperpolarised neurones, reduced the amplitudes of low threshold spikes, and diminished rebound burst firing only in WT mice. We found that 80 mg kg -1 i.p. injections of 3β-OH induced hypnosis in >60% of WT mice but failed to induce hypnosis in the majority of mutant mice. A subhypnotic dose of 3β-OH (20 mg kg -1 i.p.) accelerated induction of hypnosis by isoflurane only in WT mice, but had similar effects on the maintenance of isoflurane-induced hypnosis in both WT and Ca v 3.1 knockout mice. In vivo recordings of LFPs showed that a hypnotic dose of 3β-OH increased δ, θ, α, and β oscillations in WT mice in comparison with Ca v 3.1 knock-out mice., Conclusions: The Ca v 3.1 T-channel isoform is critical for diminished thalamocortical excitability and oscillations that underlie neurosteroid-induced hypnosis., (Copyright © 2020 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

We previously documented that the Ca V 3.3 isoform of T-type calcium channels (T-channels) is inhibited by clinically relevant concentrations of volatile anaesthetics, including isoflurane. However, little is understood about the functional role of Ca V 3.3 channels in anaesthetic-induced hypnosis and underlying neuronal oscillations. To address this issue, we used Ca V 3.3 knock-out (KO) mice and a panselective T-channel blocker 3,5-dichloro-N-[1-(2,2-dimethyltetrahydro-pyran-4-ylmethyl)-4-fluoro-piperidin-4-ylmethyl]-benzamide (TTA-P2). We found that mutant mice injected with the vehicle showed faster induction of hypnosis than wild-type (WT) mice, while the percent isoflurane at which hypnosis and immobility occurred was not different between two genotypes. Furthermore, we found that TTA-P2 facilitated isoflurane induction of hypnosis in the Ca V 3.3 KO mice more robustly than in the WT mice. Isoflurane-induced hypnosis following injections of TTA-P2 was accompanied with more prominent delta and theta EEG oscillations in the mutant mice, and reached burst-suppression pattern earlier when compared to the WT mice. Our findings point to a relatively specific value of Ca V 3.3 channels in anaesthetic induced hypnosis. Furthermore, we propose that T-channel blockers may be further explored as a valuable adjunct to reducing the usage of potent volatile anaesthetics, thereby improving their safety.

Although the central medial nucleus (CeM) of the thalamus is an essential part of the arousal system for sleep and anesthesia initiation, the precise mechanisms that regulate its activity are not well studied. We examined the role of CaV3.1 isoform of T-type calcium channels (T-channels) in the excitability and rhythmic activity of CeM neurons during isoflurane (ISO)-induced anesthesia by using mouse genetics and selective pharmacology. Patch-clamp recordings taken from acute brain slices revealed that CaV3.1 channels in CeM are inhibited by prototypical volatile anesthetic ISO (250 and 500 μM) and selective T-channels blocker 3,5-dichloro-N-[1-(2,2-dimethyl-tetrahydro-pyran-4-ylmethyl)-4-fluoro-piperidin-4-ylmethyl]-benzamide (TTA-P2). Both TTA-P2 and ISO attenuated tonic and burst firing modes, and hyperpolarized CeM neurons from wild type (WT) mice. These effects were greatly diminished or abolished in CaV3.1 null mice. Our ensuing in vivo local field potential (LFP) recordings from CeM indicated that the ability of TTA-P2 and anesthetic concentrations of ISO to promote δ oscillation was substantially weakened in CaV3.1 null mice. Furthermore, escalating ISO concentrations induced stronger burst-suppression LFP pattern in mutant than in WT mice. Our results demonstrate for the first time the importance of CaV3.1 channels in thalamocortical oscillations from the non-specific thalamic nuclei that underlie clinically important effects of ISO., (© The Author(s) 2019. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.)

Ample evidence has surfaced documenting the neurotoxic effects of various general anesthetic (GA) agents in the mammalian brain when administered at critical periods of synaptogenesis. However, little is known about how this neurotoxic insult affects persisting neuronal excitability after the initial exposure. Here we investigated synaptic activity and intrinsic excitability of the ventrobasal nucleus (VB) of the thalamus caused by neonatal GA administration. We used patch-clamp recordings from acute thalamic slices in young rats up to two weeks after neurotoxic GA exposure of isoflurane and nitrous oxide for 6 h at postnatal age of 7 (P7) days. We found that GA exposure at P7 increases evoked excitatory postsynaptic currents (eEPSCs) two fold by means through AMPA mediated mechanisms, while NMDA component was spared. In addition, miniature EPSCs showed a faster decay rate in neurons from GA treated animals when compared to sham controls. Likewise, we discovered that the amplitudes of evoked inhibitory postsynaptic currents (eIPSCs) were increased in VB neurons from GA animals about two-fold. Interestingly, these results were observed in female but not male rats. In contrast, intrinsic excitability and properties of T-type voltage gated calcium currents were minimally affected by GA exposure. Together, these data further the idea that GAs cause lasting alterations in synaptic transmission and neuronal excitability depending upon the placing and connectivity of neurons in the thalamus. Given that function of thalamocortical circuits critically depends on the delicate balance between excitation and inhibition, future development of therapies aimed at addressing consequences of altered excitability in the developing brain by GAs may be an attractive possibility., (Copyright © 2019 Elsevier Inc. All rights reserved.)