Large-conductance chloride channels of new-born rat cardiac myocytes are activated by hypotonic media

Abstract

Large-conductance chloride (LC-type Cl⁻) channel activity was studied in rat ventricular myocyte membrane during development. In contrast with results previously obtained in cultured ventricular myocytes of the new-born rat, we failed to record single-channel activity in freshly isolated myocytes whatever the age of the animal (from 2 days old to adult) and the recording patch configuration used. However, spontaneous single-channel activity of LC-type Cl⁻ channels was recorded in bleb membranes of myocytes of rats younger than 12 days, with a higher frequency in excised inside-out membrane patches than in cell-attached membrane patches. In intact neonatal myocytes, application of hypotonic media (150 mOsm) also initiated the channel activity, after variable delays (25–200 s). The channel could not be activated by suction applied through the pipette and was not observable in cells from rats older than 15 days. The LC-type Cl⁻ channels showed properties similar to those reported in other preparations and previously observed in cardiac cultured cells: they had a large single-channel conductance of 400 pS in symmetrical 150 mM NaCl, showed multiple subconductance states, a relatively high selectivity to Cl⁻ ions over Na⁺ ions (P _Cl/P _Na=24.6), were blocked by 10 μM 4-acetamido-4′-isothiocyanatostilbene-2,2′-disulphonic acid (SITS) and showed voltage-dependent inactivation. They were not activated by 10 μM colchicine or 3 μM cytochalasin D. The fact that these channels exhibit spontaneous activity when cells are swollen and membranes distended suggests that they play a role in the volume regulation of young cardiac cells, possibly through tension exerted at the channel level by cytoskeletal attachments. This channel activity is absent in adult cells.