C-type natriuretic peptide (CNP) both suppresses neurotransmitter efflux & internalizes clearance receptors

The clearance (ANP-C) receptor is perceived to function as a carrier to internalize natriuretic peptides and facilitate their destruction. However, our past data are consistent with an inhibitory neuromodulatory role for the ANP-C receptor and experiments ablating the ANP-C receptor mimicked the neuromodulatory effect of natriuretic peptides. These observations led to the following hypothesis: natriuretic peptides bind to ANP-C receptors, promoting internalization of ANP-C receptors, to reduce neurotransmitter efflux. Pheochromocytoma (PC12) cells were differentiated with Nerve Growth Factor for 7 to 10 days and then exposed to ¹²⁵I-CNP (30 pM). After 90 min at 4°C the cells were warmed to 37°C for l min and internalized radioactivity was determined. Internalized radioactivity was defined as that remaining associated with the cells after a 10 min wash with 0.2 M acetic acid. Control internalization was 28 ± 5 % per min. Staurosporine (10 nM) reduced the rate to 7 ± 5 % and ablated the neuromodulatory effect of CNP ( 100 nM). Phenylarsine oxide (20 μM) failed to prevent either internalization of CNP or neuromodulatory effects of CNP. These data are consistent with a functional role for internalization of CNP.