PACAP-induced cAMP generation is impaired in both Nf1-/- and Nf1+/- astrocytes and is independent of RAS activation. a, Nf1-/- astrocytes exhibit a marked decrease in cAMP generation in response to various concentrations of PACAP. cAMP produced by Nf1+/+ (solid line), Nf1-/- (dotted), and RAS transgenic B8 astrocytes (dashed) are shown. The inset illustrates neurofibromin and PAC-1-R expression in Nf1+/+ and Nf1-/- astrocytes after adenoviral transduction. b, Pretreatment of either Nf1+/+ (white bar) or Nf1-/- astrocytes (black bar) with the RAS farnesyltransferase inhibitor (l-744,832) did not significantly alter PACAP-mediated cAMP generation in Nf1-/- or wild-type astrocytes. The inset demonstrates that l-744,832 treatment resulted in inhibition of MAPK activity (phospho-MAPK) without altering total MAPK (MAPK) expression. c, cAMP generation in response to PACAP stimulation in also impaired in Nf1+/- astrocytes (gray bar) compared with Nf1+/+ (white bar) astrocytes.d,Whereas differences in cAMP levels were observed in Nf1+/+ (white bar) versus Nf1-/- (black bar) astrocytes in response to PACAP treatment, no differences were observed in response to EGF treatment. Each error bar represents mean ± SD. Asterisks denote statistically significant differences (p < 0.004).