INWARD CALCIUM CURRENTS IN CULTURED AND FRESHLY ISOLATED DETRUSOR MUSCLE CELLS: : EVIDENCE OF A T-TYPE CALCIUM CURRENT

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Abstract
Purpose: We carefully examined the possible routes of Ca2+ influx, and determined whether cultured cells retain Ca2+ channels and whether the culturing process changes their properties. Materials and Methods: Inward currents were measured under voltage clamp in freshly isolated cells and myocytes from confluent cell cultures of detrusor smooth muscle. Results: In guinea pig and human cells mean peak inward current density plus or minus standard deviation decreased significantly in cell culture (2.0 ± 0.9 versus 4.5 ± 2.2 pA.pF.−1) but there was no species variation. In primary cultured and passaged guinea pig cells an inward current was identified as L-type Ca2+ current. In freshly isolated cells another component to the inward current was identified that was insensitive to 20 μmol. l−1 verapamil and 20 to 50 μmol. l−1. cadmium chloride but abolished by 100 μmol. l−1 nickel chloride and identified as T-type Ca2+ current. In addition, total inward current was greater at a holding potential of −100 than −40 mV., also indicating a component of current activated at negative voltage. Steady state activation and inactivation curves of the net inward current were also compatible with a single component in cultured cells but a dual component in freshly isolated cells. The action potential was completely abolished in cultured cells by L-type Ca2+ channel blockers but incompletely so in freshly isolated cells. Outward current depended strongly on previous inward current, suggesting a predominant Ca2+ dependent outward current. Conclusions: In freshly isolated guinea pig cells T and L-type Ca2+ current is present but T-type current is absent in confluent cultures.

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