Intracellular pH dependent regulation of neuronal nitric oxide synthase activity in cultured mouse macula densa cells

Abstract

Kidney macula densa cells sense the chloride concentration of the fluid in the adjacent lumen and control glomerular filtration rate. We have used a functionally intact macula densa cell line (NE-MD) established from immortalized renal cells in culture. NE-MD cells specifically express neuronal nitric oxide synthase (nNOS) regulated by either low NaCl intake or furosemide (an inhibitor of Na⁺-K⁺-2Cl⁻ symporter). We have examined whether L-arginine-induced NO production is pH dependent or not in NE-MD cells, by using a NO-sensitive electrode. The NO production was low in control, but increased when NE-MD cells were treated with furosemide (12 µM) for 2 hrs. When furosemide-treated NE-MD cells were incubated in the presence of 100 μM amiloride (an inhibitor of Na⁺/H⁺ exchanger), L-arginine-induced NO production was unaffected. However, the NO production significantly decreased by 42% when the cells were placed in a solution containing amiloride. Similar results were obtained when NE-MD cells were incubated in the low pH solution (pH=7.1). These results strongly suggest that furosemide-induced expression of nNOS protein is not sensitive to acidosis, but its activity is pH sensitive. This may partly account for polyuria in abnormalities of electrolyte and acid-base balance. [J Physiol Sci. 2006;56 Suppl:S140]