Alterations of nPKC distribution, but normal Akt/PKB activation in denervated rat soleus muscle.
Denervation has been shown to impair the ability of insulin to stimulate glycogen synthesis and, to a lesser extent, glucose transport in rat skeletal muscle. Insulin binding to its receptor, activation of the receptor tyrosine kinase and phosphatidylinositol 3'-kinase do not appear to be involved. On the other hand, it has been shown that denervation causes an increase in the total diacylglycerol (DAG) content and membrane-associated protein kinase C (PKC) activity. In this study, we further characterize these changes in PKC and assess other possible signaling abnormalities that might be related to the decrease of glycogen synthesis. The results reveal that PKC-epsilon and -theta;, but not -alpha or -zeta, are increased in the membrane fraction 24 h after denervation and that the timing of these changes parallels the impaired ability of insulin to stimulate glycogen synthesis. At 24 h, these changes were associated with a 65% decrease in glycogen synthase (GS) activity ratio and decreased electrophoretic mobility, indicative of phosphorylation in GS in muscles incubated in the absence of insulin. Incubation of the denervated soleus with insulin for 30 min minimally increased glucose incorporation into glycogen; however, it increased GS activity threefold, to a value still less than that of control muscle, and it eliminated the gel shift. In addition, insulin increased the apparent abundance of GS kinase (GSK)-3 and protein phosphatase (PP)1 alpha in the supernatant fraction of muscle homogenate to control values, and it caused the same increases in GSK-3 and Akt/protein kinase B (PKB) phosphorylation and Akt/PKB activity that it did in nondenervated muscle. No alterations in hexokinase I or II activity were observed after denervation; however, in agreement with a previous report, glucose 6-phosphate levels were diminished in 24-h-denervated soleus, and they did not increase after insulin stimulation. These results indicate that alterations in the distribution of PKC-epsilon and -theta; accompany the impairment of glycogen synthesis in the 24-h-denervated soleus. They also indicate that the basal rate of glycogen synthesis and its stimulation by insulin in these muscles are diminished despite a normal activation of Akt/PKB and phosphorylation of GSK-3. The significance of the observed alterations to GSK-3 and PP1 alpha distribution remain to be determined.
Diabetes and Metabolism Unit and Department of Physiology, Boston University Medical Center, Boston, Massachusettes 02118, USA.
Yenshou
Y
Lin
Matthew J
MJ
Brady
Kristen
K
Wolanske
Richard
R
Holbert
Neil B
NB
Ruderman
Gordon C
GC
Yaney
D000818:Animals; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism; D004789:Enzyme Activation / Q000502:physiology; D019298:Glucose-6-Phosphate / Q000378:metabolism; D006006:Glycogen Synthase / Q000378:metabolism; D038362:Glycogen Synthase Kinase 3; D006593:Hexokinase / Q000378:metabolism; D007328:Insulin / Q000494:pharmacology; D008297:Male; D009121:Muscle Denervation; D018482:Muscle, Skeletal / Q000201:enzymology* / Q000294:innervation* / Q000378:metabolism; D010749:Phosphoprotein Phosphatases / Q000378:metabolism; D011493:Protein Kinase C / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt; D051381:Rats; D017207:Rats, Sprague-Dawley; D012016:Reference Values; D013997:Time Factors; D014018:Tissue Distribution
D007328:Insulin; D011518:Proto-Oncogene Proteins; D019298:Glucose-6-Phosphate; D006006:Glycogen Synthase; D006593:Hexokinase; C494920:Akt1 protein, rat; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt; D011493:Protein Kinase C; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D038362:Glycogen Synthase Kinase 3; D010749:Phosphoprotein Phosphatases