Forskolin, adenylyl cyclase activation

Forskolin Adenylyl cyclase activator FRTL5 rat cell line X X X... 

An inhibitory effect of 8-OH-DPAT on forskolin-stimulated adenylyl cyclase activity is observed in hippocampal membranes of the guinea pig and rat, and has been used as an index of the functional activities of 5-HTIA receptors [178]. Cold stress induces a reduction of the inhibitory effect of 8-OH-DPAT in the hippocampus isolated from 18-month-old rats, although it has no influence on either the affinity or number of [3H]8-OH-DPAT binding sites. The administration of EGb (50 mg/kg p.o. for 14 days) prevents the cold stress-induced reduction in the inhibitory effect of 8-OH-DPAT on forskolin-stimulated adenylyl cyclase activity in old rats. These results indicate that EGb prevents the stress-induced desensitization of hippocampal 5-HTu receptors thus, its effects might explain anti-stress and antidepressant properties of EGb [177]. 

Despite this A3 AR desensitisation, the adenylyl cyclase activity is not reduced, but almost a twofold increase is detected in the presence or in the absence of forskolin. This effect seems not to be mediated by e new receptor synthesis, but is probably due to an increased coupling efficiency between Gs and adenylyl cyclase. The sensitization of adenylyl cyclase activity is evidence that acute and chronic treatment of adenosine receptors might lead to opposite effects (Palmer et al. 1997). 

Lee et al. have found that 2-Ara-Gl produced a potent, dose-related inhibition of T and B cell proliferation. 2-Ara-Gl also inhibited forskolin-stimu-lated adenylyl cyclase [76], In contrast, anandamide did not inhibit forsko-lin-stimulated adenylyl cyclase activity in splenocytes, and did not inhibit T or B cell proliferation. Anandamide and 2-Ara-Gl apparently differ considerably in their biochemical profiles in the immune system. 

Fig. 8. Enhanced signaling of the a2ALys251 polymorphic receptor. Adenylyl cyclase activities were determined in the presence of 5.0 pM forskolin and the indicated concentrations of the full agonist epinephrine. Results as shown are the percentage inhibition of forskolin-stimulated activities from clones at matched levels of expression (-2500 fmol/mg)., p < 0.05 for the maximal inhibition compared with wild-type for both agonists. (From ref. 22 with permission.)...

The results of cell culture models have yielded convincing evidence demonstrating that subchronic cannabinoid administration leads to desensitization of adenylyl cyclase and ERKl/2 (Rinaldi-Carmona et al. 1998). In contrast, the results have been somewhat mixed when attempts have been made to examine this signaling pathway in whole animals. Mice treated repeatedly with CP 55,940 did not exhibit altered CBi-mediated inhibition of adenylyl cyclase in the cerebellum (Fan et al. 1996). In contrast, several other studies have reported increases in basal, forskolin or Ca -stimulated adenylyl cyclase activity in the cortex, striatum, and cerebellum of mice treated subchronically with THC (Hutcheson et al. 1998 Rubino et al. 2000b Tzavara et al. 2000). 

FIGURE 9.14 Effects of adenosine receptor agonist 2-chloro-adenosine on vascular perfusion pressure of isolated perfused rat kidneys. Minor effects seen in untreated kidneys (filled circles) and pronounced vasoconstriction while vasodilatation in kidneys coperfused with subthreshold concentrations of a-adrenoceptor vasoconstrictor methoxamine and vasodilatatory activation of adenylyl cyclase with forskolin (open circles). Redrawn from [49]. 

Adenylyl Cyclases. Figure 4 Regulation of adenylyl cyclases by G-proteins. Abbreviations Hs, Hj, Rs, and Rj denote hormones and receptors that lead to stimulation or inhibition, respectively, of adenylyl cyclases, Ca and Ci are active and inactive configurations of adenylyl cyclase, Fo forskolin binding site, Gs and Gj are GTP-dependent regulatory proteins comprising their respective as, and (3y subunits. 

Forskolin is a diteipene derivative from the plant Coleus forskohlii. It activates all mammalian isozymes of adenylyl cyclase except AC9 and AC 10. 

Active derivatives of forskolin include 7-deacetyl-forskolin (EC5o 20(llM), 6-acetyl-7-deacetyl-forskolin (EC5o 40(llIVI), 7-deacetyl-7-0-hemisuccinyl-forskolin (EC50 50(llIVI). The last of these has been used as an immobilized affinity chromatography ligand for the purification of adenylyl cyclases from tissues. 

P-site ligands inhibit adenylyl cyclases by a noncompetitive, dead-end- (post-transition-state) mechanism (cf. Fig. 6). Typically this is observed when reactions are conducted with Mn2+ or Mg2+ on forskolin- or hormone-activated adenylyl cyclases. However, under- some circumstances, uncompetitive inhibition has been noted. This is typically observed with enzyme that has been stably activated with GTPyS, with Mg2+ as cation. That this is the mechanism of P-site inhibition was most clearly demonstrated with expressed chimeric adenylyl cyclase studied by the reverse reaction. Under these conditions, inhibition by 2 -d-3 -AMP was competitive with cAMP. That is, the P-site is not a site per se, but rather an enzyme configuration and these ligands bind to the post-transition-state configuration from which product has left, but before the enzyme cycles to accept new substrate. Consequently, as post-transition-state inhibitors, P-site ligands are remarkably potent and specific inhibitors of adenylyl cyclases and have been used in many studies of tissue and cell function to suppress cAMP formation. 

Not all opiates induce an increase in forskolin stimulation of adenylyl cyclase. Buprenorphine, which is used to treat morphine dependence, does not cause a compensatory rise in cyclase activity [80]. Co-treatment of morphine with buprenorphine prevented morphine from increasing cyclase activity suggesting that bu-... 

Biochemical and molecular cloning studies indicate the existence of nine separate and unique forms of adenylyl cyclase (AC), which comprise a distinct enzyme family, referred to as AC1-AC9 [1, 2]. These members of the adenylyl cyclase superfamily are all membrane-bound. There is also an additional soluble form, sAC, that has catalytic activity similar to the others but is genetically the most divergent member of the family. All the membrane-bound forms of adenylyl cyclase are activated by the stimulatory G protein Gas (see Ch. 19), and all with the exception of AC9 are stimulated by forskolin. The soluble form sAC is not stimulated by either Gas or forskolin but is sensitive to bicarbonate levels. All known forms of... 

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