Effects Of PACAP27 On Atrial Natriuretic Peptide Secretion In Perfused Beating Rabbit Atria

Heart as an endocrine gland generating and secreting natriuretic peptides hormone.Now, natriuretic peptides hormone store in human and mammal as family of natriureticpeptides (NPs) forming, the members in NPs including ANP (atrial natriuretic peptide,ANP),BNP (brain natriuretic peptide, BNP),CNP (C-type natriuretic peptide, CNP) andDNP (dendroaspis natriuretic peptide, DNP). It is known that all of the NPs membersinvolved in important biological effects such as diuresis and natriuresis, regulating bodyfluid and blood pressure. ANP is the first member of NPs that it was synthesized andstored in atrial myocytes, it could participate in vasorelaxation, decreasing blood pressure,diuresis and natriuresis, inhibition of cellular proliferation, regulating immunization,protecting cell and lipolysis by combining with natriuretic peptide receptors which isstored in cardiovascular system, central nervous system, lymph tissue, reproductiveorgans and kidney.Atrial stretch (eg. elevate venous blood volume) is a factor by which increasesatrial secretory granule secreting of ANP. In addition, humoral factor can contribute inatrial ANP secretion. Glucocorticoid stimulate ANP secretion in vivo, angiotensinâ…¡,phentolamine, endothelin, antidiuretic hormone and prostaglandin increase plasm ANPconcentration via intravenous injection. In contrast, nitric oxide inhibits atrial ANPsecretion. Rising extracellular osmolarity and high sodium concentration or hypoxiastimulate of ANP secretion in isolated heart. But the roles of adrenergic, cholinergic andpeptidergic in the regulation of ANP release is controversial.Pituitary adenylate cyclase activating polypeptide (PACAP) was first isolated byMiyata et al. (1989) from the bovine hypothalamus on the basis to stimulate adenylatecyclase. PACAP is pleiotropic neuropeptide that belongs to the glucagons/secretine/vasoactive intestinal polypeptide (VIP)/growth hormone releasing hormone (GHRH)family. There are two bioactive forms of PACAP: PACAP38 and PACAP 27, which arewidely expressed in hypothalamus, central nervous system, peripheral nervous system and non nervous tissues, playing biological effect as neurotransmitter/modulator,especially neurotrophic factor.Previous studies showed that PACAP38 and PACAP27 have been found in the ratatria, and the PACAP receptors have been identified in the heart. At present, therelationship between atrial ANP secretion and PACAP is unclear, so purpose of thepresent study is to define the effect and the mechanism of PACAP27 on atrial ANPsecretion in perfused beating rabbit atria.The results of the present study showed that:1. PACAP27 (100 nmol/L) inhibited ANP secretion (p＜0.01) significantly withincreasing cAMP efflux (p＜0.001) and the atrial stroke volame (p＜0.01).2. PACAP6-27(1.0μmol/L), a PACAP receptor inhibitor, increased ANPsecretion (p＜0.01) significantly without changes in cAMP efflux and the atrialstroke volume. In the presence of PACAP6-27, PACAP27 (100 nmol/L)inhibited ANP secretion (p＜0.05) and increment of cAMP efflux (p＜0.001)without changes in atrial stroke volume.3. L-type Ca²⁺-channel blocker nicardipine (1.0μmol/L) significantly increasedatrial ANP secretion (P＜0.05) with decreased in atrial pulse pressure andstroke volume (P＜0.001). In the presence of nicardipine, PACAP27 (100nmol/L) increased ANP secretion (p＜0.05) concomitantly with decreased inatrial pulse pressure and stroke volume (p＜0.001).4. Non-specific protein kinases inhibitor staurosporine (1.0μmol/L) significantlyincreased atrial ANP secretion with decreased in atrial pulse pressure andstroke volume (P＜0.001). In the presence of staurosporine, atrial ANPsecretion was increased (p＜0.01) concomitantly with decreased in atrial pulsepressure and stroke volume (p＜0.001) by PACAP27 (100 nmol/L).5. H-89 (3.0μmol/L), an inhibitor of cAMP dependent protein kinase, increasedatrial ANP secretion (p＜0.05) and inhibited atrial pulse pressure and strokevolume (P＜0.05). In the presence of H-89, PACAP27 (100 nmol/L) increasedsecretion of ANP (p＜0.05) and decreased in atrial stroke volume (p＜0.05)without changes in atrial pulse pressure.6. Forskolin (1.0μmol/L), a directly activator of adenylyl cyclase stronglyinhibited ANP secretion (p＜0.01)concomitantly with increased in atrial strokevolume and cAMP efflux (p＜0.01), as well in a certain range the more production of cAMP the more inhibition of ANP secretion, and therelationship between cAMP production and ANP secretion showed inverseratio (Y=-5.268 X+9.506, r²=0.971, P＜0.001).7. L-type Ca²⁺-channel blocker nifedipine (1.0μmol/L) significantly increasedANP secretion (p＜0.01) with decreased in atrial stroke volume (P＜0.01).Whereas the nifedipine failed to modulation the inhibition of forskolin onANP secretion and the increment of cAMP production. In the presence ofnifedipine, forskolin indused-increase in atrial stroke volume was notobserved and showed decrement in atrial stroke volume (P＜0.01).8. Staurosporine (1.0μmol/L), a non-specific inhibitor of protein kinases,slightly decreased atrial stroke volume without changes in ANP secretion andcAMP effiux. Staurosporine attenuated forskolin (1.0μmol/L) induced-inhibition of ANP secretion with increased cAMP effiux (p＜0.01). In thepresence of staurosporine, forskolin induced-increase in atrial stroke volumewas not observed and showed decrement in atrial stroke volume (P＜0.01).These results indicate that:1. PACAP27 and adenylyl cyclase directly activator forskolin inhibit atrial ANPsecretion with increasing cAMP production;2. The role of PACAP27 on ANP secretion via the AC-cAMP-PKA and L-typeCa²⁺-channel signaling pathway;3. Forskolin suppress the ANP secretion via protein kinase-dependent and L-typeCa²⁺-channel-independent signaling pathway.