Contribution of reverse Na+–Ca2+ exchange to spontaneous activity in interstitial cells of Cajal in the rabbit urethra

Interstitial cells of Cajal (ICC) isolated from the rabbit urethra exhibit regular Ca 2+ oscillations that are associated with spontaneous transient inward currents (STICs) recorded under voltage clamp. Their frequency is known to be very sensitive to external Ca 2+ concentration but the mechanism of this has yet to be elucidated. In the present study experiments were performed to assess the role of Na + –Ca 2+ exchange (NCX) in this process. Membrane currents were recorded using the patch clamp technique and measurements of intracellular Ca 2+ were made using fast confocal microscopy. When reverse mode NCX was enhanced by decreasing the external Na + concentration [Na + ] o from 130 to 13 m m , the frequency of global Ca 2+ oscillations and STICs increased. Conversely, inhibition of reverse mode NCX by KB-R7943 and SEA0400 decreased the frequency of Ca 2+ oscillations and STICs. Application of caffeine (10 m m ) and noradrenaline (10 μ m ) induced transient Ca 2+ -activated chloride currents ( I ClCa ) at −60 mV due to release of Ca 2+ from ryanodine- and inositol trisphosphate (IP 3 )-sensitive Ca 2+ stores, respectively, but these responses were not blocked by KB-R7943 or SEA0400 suggesting that neither drug blocked Ca 2+ -activated chloride channels or Ca 2+ release from stores. Intact strips of rabbit urethra smooth muscle develop spontaneous myogenic tone. This tone was relaxed by application of SEA0400 in a concentration-dependent fashion. Finally, single cell RT-PCR experiments revealed that isolated ICC from the rabbit urethra only express the type 3 isoform of the Na + –Ca 2+ exchanger (NCX3). These results suggest that frequency of spontaneous activity in urethral ICC can be modulated by Ca 2+ entry via reverse NCX.