title: (R)-roscovitine prolongs the mean open time of unitary N-type calcium channel currents.
abstract: (R)-roscovitine (Ros) is a cyclin-dependent kinase inhibitor that also has been shown to have direct agonist and antagonist actions on Ca(v)2.1 (P/Q-type) and Ca(v) 2.2 (N-type) families of voltage-gated calcium channels. These kinase-independent effects represent a novel opportunity to advance our understanding of calcium channel function and calcium-triggered neurotransmitter release. Furthermore, such actions on calcium channels may direct the development of Ros derivatives as new therapeutic agents. We used patch clamp recordings to characterize mechanisms that underlie the agonist effects of Ros on unitary N-type calcium channel gating. We found that N-type channels normally gate with either a short or long mean open time, that Ros significantly prolonged the mean open time of the long gating component and increased the probability of observing channels that gated with a long open time, but had no effect on single channel conductance. Using Monte Carlo simulations of a single channel kinetic model and Ros interactions, we were able to reproduce our experimental results and investigate the model's microscopic dynamics. In particular, our simulations predicted that the longer open times generated by Ros were due to the appearance of a long open state combined with an increased amount of time spent in transitions between open states. Our results suggest a mechanism for agonist effects of Ros at the level of single channels, and provide a mechanistic explanation for previously reported agonist effects on whole cell calcium currents.
authors.affiliation: Department of Neuroscience, University of Pittsburgh, Pittsburgh, PA 15260, USA.
authors.forename: N R
authors.initials: NR
authors.lastname: DeStefino
authors.forename: A A
authors.initials: AA
authors.lastname: Pilato
authors.forename: M
authors.initials: M
authors.lastname: Dittrich
authors.forename: S V
authors.initials: SV
authors.lastname: Cherry
authors.forename: S
authors.initials: S
authors.lastname: Cho
authors.forename: J R
authors.initials: JR
authors.lastname: Stiles
authors.forename: S D
authors.initials: SD
authors.lastname: Meriney
mesh_terms: D000818:Animals; D020864:Calcium Channels, N-Type / Q000187:drug effects* / Q000378:metabolism; D020013:Calcium Signaling / Q000187:drug effects / Q000502:physiology; D002460:Cell Line; D002462:Cell Membrane / Q000187:drug effects* / Q000378:metabolism; D003198:Computer Simulation; D015640:Ion Channel Gating / Q000187:drug effects* / Q000502:physiology; D007700:Kinetics; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D009010:Monte Carlo Method; D009474:Neurons / Q000187:drug effects* / Q000378:metabolism; D018408:Patch-Clamp Techniques; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D011687:Purines / Q000494:pharmacology*; D051381:Rats; D000077546:Roscovitine; D009435:Synaptic Transmission / Q000187:drug effects / Q000502:physiology; D013997:Time Factors
chemical_list: D020864:Calcium Channels, N-Type; D047428:Protein Kinase Inhibitors; D011687:Purines; D000077546:Roscovitine