Basal dephosphorylation controls slow gating of L‐type Ca²⁺ channels in human vascular smooth muscle

Abstract

The role of cellular phosphatase activity in regulation of smooth muscle L-type Ca 2+ channels was investigated using tautomycin, a potent and specific inhibitor of serin/threonin phosphatases type 1 and 2A. Tautomycin (1-100 nM) inhibited Ca 2÷ channel activity in smooth muscle cells isolated from human umbilical vein. Tautomycin-induced inhibition of Ca 2÷ channel activity was due to a reduction of channel availability which originated mainly from prolongation of the lifetime of unavailable states of the channel. Pretreatment of smooth muscle cells with the protein kinase inhibitor H-7 (10 ttM) prevented the inhibitory effect of tautomycin. Our results suggest modulation of slow gating between available and unavailable states as a mechanism of phosphorylation-dependent down-regulation of Ca 2÷ channels in vascular smooth muscle.Key words: L-type Ca 2+ channel; Protein phosphatase; Cellular regulation; Vascular smooth muscle; Patch clamp closed states. An increase in availability as well as in open probability of available channels was found as the basis of phosphorylation-dependent up-regulation of cardiac channels [3,8]. The changes in gating behavior associated with modulatory phosphorylation of smooth muscle L-type Ca 2+ channels have not yet been clearly identified.The present study was aimed at characterization of changes in the gating behavior of single Ca 2+ channels induced by inhibition of basal dephosphorylation in vascular smooth muscle cells. Our results suggest basal activity of protein phosphatases as a major determinant of kinetic properties of L-type channels in vascular smooth muscle. We provide evidence for inhibitory modulation of slow gating between available and unavailable states as the basis of phosphorylation-dependent down-regulation of smooth muscle Ca 2÷ channels. Materials and methods

Participation of cAMP in tacrine-induced gastric smooth muscle relaxation

“…The effect is largely influenced by the type of tissues studied and the cAMP concentration; however, the precise mechanism is unclear [36]. Most researchers are of the opinion that following phosphorylation of the Са 2+ channel proteins by PKA [36][37][38] an inhibition of the currents through them is recorded -an effect similar to that observed in our experiments.…”

Functional impairment of renal afferent arteriolar voltage-gated calcium channels in rats with diabetes mellitus.

“…The ability to rapidly restore normal responsiveness through acute alterations in extracellular glucose concentration suggests that the defect reflects an altered regulation of channel activity. Changes in basal dephosphorylation represent a major determinant of L-type VGCC kinetics in VSM cells (53,54) and likely underlie the reported effect of protein kinase C to reduce the ability of VGCCs to open upon depolarization (55). Hence, glucose-induced activation of protein kinase C in VSM (56) could suppress VGCC activity in IDDM through this type of phosphorylation-dependent process.…”

Role of serine-threonine phosphoprotein phosphatases in smooth muscle contractility

“…In mouse colon, PP1 inhibits the A-type K ϩ channel 19-pS through its fast inactivation mechanism (7,236). The human umbilical vein L-type Ca 2ϩ channel was reported to be activated by PP1 (74). However, this study conflicted with a follow-up study in the same cell type (75).…”