Endocrine Abstracts

Current therapeutics target cGMP in treating erectile dysfunction and other cardiovascular disorders. In the endocrine system, cGMP mediates the vast majority of biological effects of the natriuretic peptides (ANP, BNP and CNP) acting via their specific guanylyl cyclases receptors (GC-A and GC-B). There are severe endocrine consequences for mice and humans exhibiting mutations to GC-B receptors, including achondroplasia, growth hormone deficiency and female infertility. Despite this, pharmacological regulation of GC-B in endocrine tissues is poorly established. In this study, we used the rat somatolactotroph GH3 cell line, which expresses endogenous GC-B receptors, to examine GC-B desensitisation and down-regulation. cGMP accumulation (measured by cGMP-EIA; samples generated in the presence of 1 mM IBMX) in response to CNP (100 nM) was rapid and significant between 0 and 5 min (r²=0.85, ***P<0.001), yet this accumulation was not maintained by 15 min (r²=0.32, P>0.5). As receptor dephosphorylation is thought to mediate GC-B desensitisation, similar studies were performed except using 30 min pre-treatments of phosphatase inhibitors (okadaic acid, cypermethrin, sodium orthovanadate) prior to a short time-course exposure to CNP (0, 5, 15 min) in the presence of IBMX. Both okadaic acid and cypermethrin partially rescued GC-B signalling, as indicated by a significant rate of cGMP accumulation between 5 and 15 min (r²=0.69, *P<0.05), suggesting a potential role for protein phosphatases 1 and 2 in GC-B desensitisation. However, sodium orthovanadate did not rescue GC-B signalling and inhibited total cGMP accumulation suggesting blockade of tyrosine dephosphorylation interferes with GC-B signalling. Finally, internalisation experiments (using confocal microscopy) revealed that the GC-B receptor did not internalise following a short or long exposure to CNP. Collectively, these data suggest pituitary GC-B signalling is mediated by protein phosphatases 1 and 2, but not receptor internalisation. These data may inform the design of potential treatments for GH deficiency with CNP analogues.