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Phorbol Ester‐Mediated Stimulation of Phospholipase D Activity in Sciatic Nerve from Normal and Diabetic Rats
Author(s) -
Eggen Bart J. L.,
Eichberg Joseph
Publication year - 1992
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1111/j.1471-4159.1992.tb08462.x
Subject(s) - phosphatidylethanol , protein kinase c , phospholipase c , medicine , endocrinology , phospholipid , phorbol , phospholipase , phospholipase d , phosphatidylcholine , chemistry , stimulation , sciatic nerve , biology , enzyme , biochemistry , membrane
Evidence for the presence of phospholipase D activity in sciatic nerve was obtained by incubation of 32 P‐prelabeled nerve segments in the presence of ethanol and measurement of [ 32 P]phosphatidylethanol (PEth) formation expressed as a fraction of total phospholipid radioactivity. PEth synthesis was enhanced with increasing concentrations of ethanol (100 m M ‐2 M ). 4‐β‐Phorbol dibutyrate (100 n M‐ 1 μM ) stimulated PEth formation up to twofold in a time‐ and dose‐dependent manner. The stimulatory effect evoked by 100 n M phorbol ester was completely abolished by Ro 31–8220 (compound 3), a selective protein kinase C inhibitor. Efforts to identify the phospholipid precursor of PEth were unsuccessful, suggesting this product arises from a small discrete precursor pool. On subcellular fractionation of nerve, the ratio of basal and 4‐β‐phorbol dibutyrate‐stimulated phospholipase D activity recovered in a myelin‐enriched fraction, compared with a nonmyelin fraction, was 0.5 when results are expressed as a percentage of total phospholipid radioactivity. This ratio rises to 1.2 if the results are calculated assuming only phosphatidylcholine and phosphatidylethanolamine are potential precursors. The results suggest that myelin is a major locus of phospholipase D activity. Nerve from streptozotocin‐induced diabetic and control animals displayed the same basal phospholipase D activity, but the enzyme in diabetic nerve was stimulated to a greater extent by a suboptimal concentration of 4‐β‐phorbol dibutyrate. These results support the conclusion that protein kinase C modulates phospholipase D activity in nerve and suggest that in diabetic nerve the enzyme activation mechanism may possess increased sensitivity.