Study of nonspecific cation channel coupled to P2z purinergic receptors using an acid load technique

The intracellular pH (pHi) of rat submandibular cells was measured by 2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein (BCECF). The cells recovered from ammonium (30 mM) prepulse to their resting pHi within 10 min. Ethylisopropylamiloride (EIPA), an inhibitor of the Na+/H+ exchanger, slows the rate of pHi recovery. ATP (1 mM), in the presence of EIPA, increases the rate of recovery 3.7-fold in the absence or presence of ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid. The recovery was blocked by the addition of 5 mM Mg2+ or 10 microM Coomassie blue. The response was elicited by 2'- and 3'-O-(4-benzoylbenzoyl)-adenosine 5'-triphosphate but not by ADP, UTP, adenyl (beta-gamma-methylene)-diphosphonate, 2-methylthioadenosine 5'-triphosphate, or muscarinic or beta-adrenergic agonists. The purinergic response was also observed when the cells were acidified by sodium propionate and could not be mimicked by the depolarization of the plasma membrane. Aluminum fluoride did not reproduce the response to ATP, suggesting that the observed response does not involve a high-molecular-weight GTP-binding protein. It is concluded that the activation of P2z receptors, probably by the opening of nonspecific cation channels, increases the permeability to protons in rat submandibular glands.