In skeletal muscle, a local increase of acetylcholine (ACh) in a few endplates has been hypothesized to cause the formation of contraction knots that can be found in myofascial trigger points. To test this hypothesis, in rats, small amounts of an acetylcholinesterase inhibitor (diisopropylfluorophosphate, DFP) were injected into the proximal half of the gastrocnemius muscle, and the muscle nerve was electrically stimulated for 30-60 min for induction of muscle twitches. The distal half of the muscle, served as a control to assess the effects of the twitches without DFP. Sections of the muscle were evaluated for morphological changes in relation to the location of blocked endplates. In comparison to the distal half of the muscle, the DFP-injected proximal half exhibited significantly higher numbers of abnormally contracted fibers (local contractures), torn fibers, and longitudinal stripes. DFP-injected animals in which the muscle nerve was not stimulated, and which were allowed to survive for 24 h, exhibited the same lesions but in smaller numbers. The results support the assumption that a dysfunctional endplate exhibiting increased release of ACh may be the starting point for regional abnormal contractions which are thought to be essential for the formation of myofascial trigger points.