Hydrolysis of the Cyclic 3′:5′‐Monophosphates of Adenosine and Guanosine by Rat Pancreas

Abstract

I. Crude rat pancreas homogenates had two apparent Km values for hydrolysis of adenosine 3':5'-monophosphate (cyclic AMP) of 1.9 and 32 pM with corresponding apparent V values of 147 and 1033 pmol x min-l x mg protein-l a t 30 "C. The same homogenates exhibited two apparent K, for hydrolysis of guanosine 3':5'-monophosphate (cyclic GMP) of 4 and 40 pM with apparent V values of 70 and 300 pmol x rnin-l x mg protein-I, respectively. I n comparison with liver and parotids the affinity of low-Km phosphodiesterases for cyclic nucleotides was low but the velocity of hydrolysis was higher. Cyclic GMP interfered somewhat with cyclic AMP hydrolysis but it was not possible to define Ki values. Aminophylline was a weak non-competititive inhibitor of both forms of cyclic AMP phosphodiesterases.2. A major portion (6501,) of both cyclic AMP phosphodiesterase activities and of the enzyme with a 40-pM K m €or cyclic GMP was confined to the cytosol while the remnant was partitioned among particulate fractions. The subcellular distribution of the enzyme, presenting a 4-pM Km for cyclic GMP, differred since as much as 82O/, of its activity was recovered in the cytosol.3. A secretory cycle induced in vivo by pilocarpine (30 mg/kg) did not influence phosphodiesterase activities for 3 h after the intraperitoneal injection. A 10-day adaptation to protein malnutritution tended to increase the activity of the low-Km cyclic AMP phosphodiesterase.4. It was concluded that the rat pancreas was able to hydrolyse cyclic nucleotides a t physiological concentrations and that more than one enzyme was apparently involved.Evidence for participation of adenosine 3' : 5'-monophosphate (cyclic AMP) in stimulus-secretion coupling in the exocrine pancreas is increasing rapidly. However the data are ambiguous and the physiological effect of pancreozymin and acetylcholine on adenylate cyclase is not yet definitely clarified.The dibutyryl derivative of cyclic AMP stimulates protein secretion from isolated mouse [I] and rabbit [2,3] pancreas but not from rat [4,5] and cat [6] pancreas. I n response to pancreozymin stimulation, cyclic AMP levels do not increase in guinea-pig pancreas in vitro [7] while the rise observed in vivo in the cat [S] is not clearly associated with protein release. Adenylate cyclase in a fresh 4000 x g pellet from rat pancreas is half-maximally stimulated by a concentration of purified pancreozymin [9] exceeding 200 times the maximal concentration which is effective on intact cells. These data suggest that stimulus-secretion coupling may involve a coordinated monitoring of both adenylate cyclase and cyclic AMP phosphodiesterase(s). Over 10 years ago, Butcher and Sutherland [lo] noticed the presence of an active high-Km cyclic AMP phosphodiesterase in dog pancreas. For the last 3 years, Beavo et al. [l 11 have initiated more interest in the hydrolysis of cyclic nucleotides, with concentrations closer to physiological levels and recently Uzunov and Weiss [12] were able to differentiate up to six different activities in the rat cere...

In vivo effects of pancreozymin, secretin, vasoactive intestinal polypeptide and pilocarpine on the levels of cyclic AMP and cyclic GMP in the rat pancreas

“…A pancreatic adepylatecyclase can be stimulated by pancreozymin [5,7,8] yet the status of cyclic AMP as a second messenger of the secretagogues is ambiguous. The occurrence in the rat pancreas of phosphodiesterase activity capable of hydrolyzing physiological concentrations of cyclic GMP [9] and of cyclic GMP-dependent protein kinase [lo] suggests that cyclic GMP may intervene in pancreatic secretion. It was decided therefore to compare the in vivo time course of cyclic AMP and cyclic GMP levels following administration of pancreozymin, pilocarpine, secretin and vasoactive intestinal polypeptide (VIP).…”

Plasma cyclic AMP levels during a secretin-caerulein pancreatic function test in liver and pancreatic disease.

“…For instance, in the pancreas, a complex cyclic nucleotide phosphodiesterase system is able rapidly to hydrolyse cyclic AMP (Robberecht et al, 1974), but some cyclic AMP also diffuses into the blood (Robberecht, 1976) or escapes into the pancreatic juice (Domschke et al, 1976). A similar cyclic nucleotide disposal has been described for the liver after hormonal stimulation (Broadus et al, 1970;Levine and Hall, 1976).…”

Action of Secretin and Vasoactive Intestinal Peptide (Vip) on the Adenylate Cyclase System of Pancreatic Acinar Cells

“…Cyclic Nucleotide Phosphodiesterases. Kinetic évidence already suggests that more than one enzyme is involved in the hydrolysis of cyclic nucleotides in the rat pancréas(24,25). When the 105,000 x g supernatant is subraitted to gel filtration on Sephadex G-20G in the présence of EGTA, 3 peaks of phosphodiesterase activity are separated (26).…”

“…In the présence of 0,2 mM EGTA ail three fractions are active on low concentrations of cyclic AMP and cyclic GMP, with a préférence for the former substrate. Only the third phosphodiesterase fraction proves to have a greater affinity for cyclic GMP when activated by the pancreatic calciura-dependent activator protein(25).INTERACTION WITH OTHER GASTROINTESTINAL HORMONESOne of the major effect of secretin and VIP on the exocrine pancréas raight be to modulate the action of other gastrointestinal hormones. It is generally accepted that two partly independent séquences of events between stimulus and sécrétion can implement the same final response.…”