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Initial biochemical and functional characterization of cyclic nucleotide phosphodiesterase isozymes in canine colonic smooth muscle.
J Pharmacol Exp Ther. 1993 Feb; 264(2):801-12.JP

Abstract

Cyclic nucleotides mediate relaxation of gastrointestinal smooth muscle. The intracellular concentration of these second messengers is determined by a balance between their synthesis and metabolism. Because cyclic nucleotide phosphodiesterase enzymes (PDE) are the sole enzymes responsible for their degradation, it is essential to determine the role of the various PDE isozymes in regulating cyclic nucleotide content of gastrointestinal smooth muscle. To examine the role of different PDE isozymes in colonic smooth muscle motility, soluble PDE activity was measured in fractions obtained from homogenates of canine colon using DEAE sepharose chromatography. PDE activity was determined using [3H]cyclic AMP (cAMP) (1 microM) or [3H]cyclic GMP (cGMP) (1 microM) as a substrate. Results indicated that colonic smooth muscle contains at least two forms of PDE with a high affinity for cGMP. One form was stimulated by calmodulin (type I) and the other was inhibited by low concentrations of zaprinast (type V). In addition, colonic smooth muscle contains at least two isozymes that prefer cAMP as a substrate. One form was inhibited by SB 94120 and cGMP (type III) and the other by rolipram (type IV). An additional peak of PDE activity was identified. The hydrolysis of cAMP by this peak was greatly enhanced by the presence of cGMP, suggesting that this activity belonged to type II or cGMP-stimulated PDE. The functional role of these isozymes was evaluated by determining the ability of selective PDE inhibitors to antagonize a carbachol (0.3 microM)-induced contraction of isolated circular colonic muscle strips in the presence of forskolin (0.2 microM). Concentration-dependent decreases in contractile activity were observed with the following potency order: rolipram > Ro 20-1724 > isobutyl methylxanthine > SB 94120 > zaprinast. These results demonstrate that colonic smooth muscle contains several PDE isozymes and that selective inhibition of PDE isozymes can increase cyclic nucleotide content and antagonize contractile responses. Functionally, PDE IV appears to be very important in reducing contractile activity, suggesting that selective PDE IV inhibitors might be useful in the treatment of gut hypermotility disorders.

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Authors+Show Affiliations

Barnette MS
Division of Pharmacology, SmithKline Beecham Pharmaceuticals, King of Prussia, Pennsylvania.
Manning CD
No affiliation info available
Price WJ
No affiliation info available
Barone FC
No affiliation info available

MeSH

1-Methyl-3-isobutylxanthine2',3'-Cyclic-Nucleotide PhosphodiesterasesAnimalsColforsinColonCyclic AMPCyclic GMPDogsIsoenzymesMuscle, SmoothPhosphodiesterase InhibitorsPurinonesPyrrolidinonesRolipram

Pub Type(s)

Journal Article

Language

eng

PubMed ID

7679736

Citation

Barnette, M S., et al. "Initial Biochemical and Functional Characterization of Cyclic Nucleotide Phosphodiesterase Isozymes in Canine Colonic Smooth Muscle." The Journal of Pharmacology and Experimental Therapeutics, vol. 264, no. 2, 1993, pp. 801-12.
Barnette MS, Manning CD, Price WJ, et al. Initial biochemical and functional characterization of cyclic nucleotide phosphodiesterase isozymes in canine colonic smooth muscle. J Pharmacol Exp Ther. 1993;264(2):801-12.
Barnette, M. S., Manning, C. D., Price, W. J., & Barone, F. C. (1993). Initial biochemical and functional characterization of cyclic nucleotide phosphodiesterase isozymes in canine colonic smooth muscle. The Journal of Pharmacology and Experimental Therapeutics, 264(2), 801-12.
Barnette MS, et al. Initial Biochemical and Functional Characterization of Cyclic Nucleotide Phosphodiesterase Isozymes in Canine Colonic Smooth Muscle. J Pharmacol Exp Ther. 1993;264(2):801-12. PubMed PMID: 7679736.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Initial biochemical and functional characterization of cyclic nucleotide phosphodiesterase isozymes in canine colonic smooth muscle. AU - Barnette,M S, AU - Manning,C D, AU - Price,W J, AU - Barone,F C, PY - 1993/2/1/pubmed PY - 1993/2/1/medline PY - 1993/2/1/entrez SP - 801 EP - 12 JF - The Journal of pharmacology and experimental therapeutics JO - J Pharmacol Exp Ther VL - 264 IS - 2 N2 - Cyclic nucleotides mediate relaxation of gastrointestinal smooth muscle. The intracellular concentration of these second messengers is determined by a balance between their synthesis and metabolism. Because cyclic nucleotide phosphodiesterase enzymes (PDE) are the sole enzymes responsible for their degradation, it is essential to determine the role of the various PDE isozymes in regulating cyclic nucleotide content of gastrointestinal smooth muscle. To examine the role of different PDE isozymes in colonic smooth muscle motility, soluble PDE activity was measured in fractions obtained from homogenates of canine colon using DEAE sepharose chromatography. PDE activity was determined using [3H]cyclic AMP (cAMP) (1 microM) or [3H]cyclic GMP (cGMP) (1 microM) as a substrate. Results indicated that colonic smooth muscle contains at least two forms of PDE with a high affinity for cGMP. One form was stimulated by calmodulin (type I) and the other was inhibited by low concentrations of zaprinast (type V). In addition, colonic smooth muscle contains at least two isozymes that prefer cAMP as a substrate. One form was inhibited by SB 94120 and cGMP (type III) and the other by rolipram (type IV). An additional peak of PDE activity was identified. The hydrolysis of cAMP by this peak was greatly enhanced by the presence of cGMP, suggesting that this activity belonged to type II or cGMP-stimulated PDE. The functional role of these isozymes was evaluated by determining the ability of selective PDE inhibitors to antagonize a carbachol (0.3 microM)-induced contraction of isolated circular colonic muscle strips in the presence of forskolin (0.2 microM). Concentration-dependent decreases in contractile activity were observed with the following potency order: rolipram > Ro 20-1724 > isobutyl methylxanthine > SB 94120 > zaprinast. These results demonstrate that colonic smooth muscle contains several PDE isozymes and that selective inhibition of PDE isozymes can increase cyclic nucleotide content and antagonize contractile responses. Functionally, PDE IV appears to be very important in reducing contractile activity, suggesting that selective PDE IV inhibitors might be useful in the treatment of gut hypermotility disorders. SN - 0022-3565 UR - https://www.unboundmedicine.com/medline/citation/7679736/Initial_biochemical_and_functional_characterization_of_cyclic_nucleotide_phosphodiesterase_isozymes_in_canine_colonic_smooth_muscle_ L2 - https://jpet.aspetjournals.org/cgi/pmidlookup?view=long&pmid=7679736 DB - PRIME DP - Unbound Medicine ER -