Enhanced capacitative calcium entry and TRPC channel gene expression in human LES smooth muscle

Transient receptor potential channel ( TRPC) genes encode Ca2+-permeable channels mediating capacitative Ca2+ entry ( CCE), which maintains intracellular Ca2+ stores. We compared TRPC gene expression and CCE in human esophageal body (EB) and lower esophageal sphincter (LES), because these smooth muscles have distinct contractile functions that are likely associated with different Ca2+ regulatory mechanisms. Circular layer smooth muscle cells were grown in primary culture. Transcriptional expression of TRPC genes was compared by semiquantitative RT-PCR. CCE was measured by fura 2 Ca2+ fluorescence after blockade of sarcoplasmic reticulum Ca2+-ATPase with thapsigargin. mRNA for TRPC1, TRPC3, TRPC4, TRPC5, and TRPC6 was identified in EB and LES. TRPC3 and TRPC4 were more abundant in LES than EB. Basal concentration of free intracellular Ca2+ ([Ca2+](i)) was similar in cells from LES (138 +/- 8 nmol/l) and EB (110 +/- 6 nmol/l) and increased with ACh (10 mumol/l; 650 +/- 28 and 590 +/- 21 nmol/l, respectively). With zero Ca2+ in bath, thapsigargin ( 2 mumol/l) increased [Ca2+](i) more in LES (550 +/- 22 nmol/l) than EB (250 +/- 15 nmol/l, P < 0.001). Subsequent external application of 1 mmol/l Ca2+ increased [Ca2+](i) more in LES (585 +/- 35 nmol/l) than EB (295 +/- 21 nmol/l, P < 0.001), indicating enhanced CCE in LES. This demonstrates CCE and TRPC transcriptional expression in human esophageal smooth muscle. In LES cells, enhanced CCE and expression of TRPC3 and TRPC4 may contribute to the physiological characteristics that distinguish LES from EB.