Natriuretic peptides (NPs), ANP, BNP and CNP display hypotensive effects in the mammalian eye by lowering the intraocular pressure (IOP), a function that is mediated by the bilayer ocular ciliary epithelium (CE), in conjunction with the trabecular meshwork. ANP regulates Na⁺/H ⁺-exchanger (NHE) activity, and inhibitors of NHE have been shown to lower IOP. In the present work, we examined whether NPs influence the NHE activity of the CE, which is comprised of PE and NPE cells, by directly recording the rate of intracellular pH (pH_i) recovery from its inner NPE cell layer. NPs inhibited, in a dose-dependent manner (1 to 100nM), the rate of pHi recovery with an order of potency CNP>>ANP>BNP indicative that this inhibition is mediated by the presence of NPR type B receptors. 8-Br-cGMP a non hydrolyzable analogue of cGMP mimicked NPs in inhibiting the rate of Na ⁺-dependent pH_i recovery. In contrast, ethylisopropylamiloride (EIPA) at 100nM or amiloride (10µM) completely abolished the pH_i recovery by NHE. 18-glycerrhitinic acid, a gap junction blocker, attenuated the inhibitory effect of CNP on the rate of pHi recovery, suggesting that the NHE activity in both cell layers of the CE is co-regulated. This interpretation was supported, in part, by the co-expression of the NHE-1 isoform mRNA in both NPE and PE cells. The mechanism by which the inhibitory effect of NPs on NHE-1 activity might influence the net solute movement or fluid transport by the bilayer CE remains to be determined.