Domain III regulates N-type (Ca_V2.2) calcium channel closing kinetics

Abstract

Yarotskyy V, Gao G, Peterson BZ, Elmslie KS. Domain III regulates N-type (Ca V 2.2) calcium channel closing kinetics. J Neurophysiol 107: 1942-1951, 2012. First published December 28, 2011 doi:10.1152 doi:10. /jn.00993.2011 and Ca V 1.2 (L-type) calcium channels gate differently in response to membrane depolarization, which is critical to the unique physiological functions mediated by these channels. We wondered if the source for these differences could be identified. As a first step, we examined the effect of domain exchange between N-type and L-type channels on activationdeactivation kinetics, which were significantly different between these channels. Kinetic analysis of chimeric channels revealed N-channellike deactivation for all chimeric channels containing N-channel domain III, while activation appeared to be a more distributed function across domains. This led us to hypothesize that domain III was an important regulator of N-channel closing. This idea was further examined with R-roscovitine, which is a trisubstituted purine that slows N-channel deactivation by exclusively binding to activated N-channels. L-channels lack this response to roscovitine, which allowed us to use N-L chimeras to test the role of domain III in roscovitine modulation of N-channel deactivation. In support of our hypothesis, all chimeric channels containing the N-channel domain III responded to roscovitine with slowed deactivation, while those chimeric channels with L-channel domain III did not. Thus a combination of kinetic and pharmacological evidence supports the hypothesis that domain III is an important regulator of N-channel closing. Our results support specialization of gating functions among calcium channel domains.

Cloning, synthesis, and characterization of αO-conotoxin GeXIVA, a potent α9α10 nicotinic acetylcholine receptor antagonist

“…The pipette solution contained 104 mM NMG·Cl, 14 mM creatine·PO 4 , 6 mM MgCl 2 , 10 mM NMG·Hepes, 5 mM Tris·ATP, 10 mM NMG 2 ·EGTA, and 0.3 Tris 2 ·GTP, with pH 7.4 and an osmolarity of 300 mOsm. Ionic currents were recorded and analyzed as previously described (19,58). Recordings were carried out at room temperature, and the holding potential was −80 mV.…”

Regulation of Neuronal Cav3.1 Channels by Cyclin-Dependent Kinase 5 (Cdk5)

“…It should be noted here, that the Cdk5 inhibitor roscovitine (Ro) has also shown to affect Ca V 3.1 channel activity. Specifically, the drug seems to inhibit these channels in part by stabilizing the closed-inactivated state [ 27 ]. In addition, it has been reported that Ro affects Ca V 2.2 (N-type) current kinetics [ 28 ].…”

New Cav2 calcium channel gating modifiers with agonist activity and therapeutic potential to treat neuromuscular disease

“…If prior use of ( R )-roscovitine by experimentalists is any indication, these novel analogs with greater potency and efficacy should be valuable tool compounds. Such tool use has previously aided the study of Cav2 single channel conductance (Weber et al, 2010), Cav2 VGCC structure and function (Yarotskyy et al, 2012), Cav2 VGCC permeation (Buraei et al, 2014), the calcium ion control of transmitter release from synapses (Wen et al, 2013; Satake and Imota, 2014), and calcium ion dynamics in non-neuronal cells (Tamma et al, 2013).…”