alpha-Melanocyte-stimulating hormone modulates nitric oxide production in melanocytes

We have previously observed that melanocytes produce nitric oxide in respon se to ultraviolet radiation and lipopolysaccharide and in this study have e xamined how these responses are affected by alpha-melanocyte-stimulating ho rmone. Nitric oxide production by cultured cells was measured electrochemic ally in real time using an ISO-nitric oxide sensor probe. B16 mouse melanom a cells released nitric oxide in response to lipopolysaccharide and the eff ects were enhanced in cells that had been grown in the presence of 10(-11)- 10(-9) M alpha-melanocyte-stimulating hormone prior to stimulation. At conc entrations in excess of 10(-9) M alpha-melanocyte-stimulating hormone decre ased nitric oxide production. Preincubation with lipopolysaccharide, a well -known inducer of inducible nitric oxide synthase, also increased nitric ox ide production but this response was reduced by alpha-melanocyte-stimulatin g hormone. alpha-Melanocyte-stimulating hormone also increased the levels o f nitric oxide produced in response to ultraviolet radiation (20-100 mJ per cm(2)) in B16 cells. The same effect was seen in human melanocytes and as this was inhibited by aminoguanidine would appear to involve an induction o f inducible nitric oxide synthase. Reverse transcription-polymerase chain r eaction showed that melanocytic cells express inducible nitric oxide syntha se mRNA. Western blotting analysis and immunocytochemistry confirmed the pr esence of inducible nitric oxide synthase protein in B16 cells and FM55 hum an melanoma cells and that the levels were increased in response to alpha-m elanocyte-stimulating hormone. alpha-Melanocyte-stimulating hormone, howeve r, decreased inducible nitric oxide synthase protein expression, which occu rred in response to lipopolysaccharide. These results suggest that alpha-me lanocyte-stimulating hormone regulates nitric oxide production in melanocyt ic cells by modulating the induction of inducible nitric oxide synthase. Ad ditional experiments showed that nitric oxide increased melanin production by B16 cells and human melanocytes. This is in keeping with a melanogenic r ole for nitric oxide but whether its production by melanocytes in response to alpha-melanocyte-stimulating hormone is associated with such a role or w hether it has some other significance relating to melanocyte differentiatio n or in mediating immunomodulatory actions of alpha-melanocyte-stimulating hormone remains to be seen.