Ca²⁺ images and K⁺ current during depolarization in smooth muscle cells of the guinea‐pig vas deferens and urinary bladder

Abstract

Electrical events and intracellular calcium concentration ([Ca2+]) imaged using fluo‐3 and laser scanning confocal microscopy were simultaneously monitored in single smooth muscle cells freshly isolated from guinea‐pig vas deferens or urinary bladder. Images obtained every 8 ms, during stepping from ‐60 to 0 or +10 mV for 50 ms under voltage clamp, showed that a rise in [Ca2+] could be detected within 20 ms of depolarization in five to twenty small (< 2 μm diameter) ‘hot spots’, over 95 % of which were located within 1.5 μm of the cell membrane. Depolarization at 30 s intervals activated hot spots at the same places. Cd2+ or verapamil abolished both hot spots and Ca2+‐activated K+ current (IK,Ca). Caffeine almost abolished hot spots and markedly reduced IK,Ca. Cyclopiazonic acid, which raised basal global [Ca2+], decreased the rise in hot spot [Ca2+] and IK,Ca amplitude during depolarization. These results suggest that Ca2+ entry caused Ca2+‐induced Ca2+ release (CICR). Under voltage clamp, hot spot [Ca2+] closely paralleled the rise in IK,Ca and reached a peak within 20 ms of the start of depolarization, but the rise in global [Ca2+] over the whole cell area was much slower. Step depolarization to potentials positive to ‐20 mV caused hot spots to grow in size and coalesce, leading to a rise in global [Ca2+] and contraction. Ca2+ hot spots also occurred during the up‐stroke of an evoked action potential under current clamp. It is concluded that the entry of Ca2+ in the early stages of an action potential evokes CICR from discrete subplasmalemma Ca2+ storage sites and generates hot spots that spread to initiate a contraction. The activation of Ca2+‐dependent K+ channels in the plasmalemma over hot spots initiates IK,Ca and action potential repolarization.

Overactive Bladder and Incontinence in the Absence of the BK Large Conductance Ca2+-activated K+ Channel

“…We observed local transient increases in Ca 2ϩ in UBSM cells from both Slo ϩ/ϩ and Slo Ϫ/Ϫ mice (Fig. 5, B and C), with spatial and temporal characteristics consistent with calcium sparks described previously (10,17,29,30,32). Furthermore, Ca 2ϩ spark amplitude and frequency were not different between Slo ϩ/ϩ and Slo Ϫ/Ϫ UBSM cells (Fig.…”

Methyl-β-cyclodextrin Prevents Ca2+-Induced Ca2+ Release in Smooth Muscle Cells of Mouse Urinary Bladder

“…A large outward current followed the initial inward current (Ca 2+ current). Most of the outward current recorded at 0 mV (by over 80%) was inhibited by addition of 1 µM paxilline or 100 nM iberiotoxin (not shown), indicating that a large part of the outward current is the BK channel current (11,13). The activation of outward current appeared to be slower in MβCD-pretreated myocytes.…”

Molecular mechanisms underlying the effect of the novel BK channel opener GoSlo: Involvement of the S4/S5 linker and the S6 segment

“…BK channels play a number of important roles that govern the excitability of neuronal and smooth muscle cells. In bladder smooth muscle, for example, they contribute significantly to the repolarization phase of the action potential and thus modulate the contractile activity of this tissue (19). Interestingly, BKα knockout mice (20) display a functionally incontinent phenotype, presumably due to detrusor overactivity.…”