A model for electrostimulation causing contractions of primary cultures of rat myotubes was established. The kinetics of glycogen degradation was investigated for a 2-h period to elucidate the coupling between contraction and glycogenolytic flux. Electrostimulation caused contraction and increased glycogenolytic flux, but had no effect on glycogen phosphorylase-a activity. Forskolin increased glycogenolytic flux more than electrostimulation, and caused a fast activation of glycogen phosphorylase, while it did not elicit contraction. The effects of electrostimulation and forskolin on glycogenolytic flux were partly additive. The metabolism of glucose and glycogen was almost equally anaerobic and aerobic. The ATP content remained constant during glycogenolysis, but phosphocreatine decreased with the largest decrease in electrostimulated cells. The calculated ATP turnover rate increased about 3 times by electrostimulation. For all conditions, pHi decreased from about 7.0 to about 6.6 at 2 h. It is concluded that in the present in vitro system glycogenolytic flux may be enhanced without eliciting contraction, a condition normally not observed in vivo. The system also shows much less dynamic range of energy metabolism than in vivo, primarily because of a high resting ATP turnover.