Sodium nitrate decreases agrin-induced acetylcholine receptor clustering.

Background: Humans are exposed to nitrate predominantly through diet with peak plasma concentrations within an hour after ingestion, but additional exposure is obtained from the environment, and minimally through de novo synthesis. Higher nitrate consumption has been associated with methemoglobinemia, spontaneous abortions, atherosclerosis, myocardial ischemia, septic and distressed lung, inflammatory bowel disease, amyotrophic lateral sclerosis, and neural tube defects. However, skeletal muscle development has not been examined.

Ethanol decreases agrin-induced acetylcholine receptor clustering in C2C12 myotube culture.

We investigated the effect of ethanol on skeletal muscle development using C2C12 cell culture. The ethanol concentrations of 10mM, 25mM, and 100mM, were tested because plasma samples of alcohol-dependent individuals fall within this range. We assessed two specific events in skeletal muscle development, the fusion of myoblasts to form myotubes and the acetylcholine receptor (AChR) clustering associated with neuromuscular synapse formation.

Caffeine and nicotine decrease acetylcholine receptor clustering in C2C12 myotube culture.

As motor neurons approach skeletal muscle during development, agrin is released and induces acetylcholine receptor (AChR) clustering. Our laboratory investigates the effect of environmental agents on skeletal muscle development by using C2C12 cell culture. For the current project, we investigated both short-term and long-term exposure to caffeine, nicotine, or both, at physiologically relevant concentrations.

The pesticide methoxychlor decreases myotube formation in cell culture by slowing myoblast proliferation.

We studied the effect of the estrogenic pesticide methoxychlor (MXC) on skeletal muscle development using C2C12 cell culture. Myoblast cultures were exposed to various concentrations of MXC at various times during the process of myoblast fusion into myotubes. We observed that MXC exposure decreased myotube formation.

Reduced glycosaminoglycan sulfation diminishes the agrin signal transduction pathway.

Proteoglycans consist of a protein core complexed to glycosaminoglycan (GAG) side chains, are abundant in skeletal muscle cell membranes and basal lamina, and have important functions in neuromuscular synapse development. Treatment with chlorate results in the undersulfation of heparan sulfate and chondroitin sulfate GAGs in cell culture. In addition, chlorate treatment decreases the frequency of spontaneous acetylcholine receptor (AChR) clustering in skeletal muscle cell culture.

Nicotine decreases agrin signaling and acetylcholine receptor clustering in C2C12 myotube culture.

The clustering of acetylcholine receptors (AChRs) in skeletal muscle fibers is a critical event in neuromuscular synaptogenesis. AChRs in concert with other molecules form postsynaptic scaffolds in response to agrin released from motor neurons as motor neurons near skeletal muscle fibers in development. Agrin drives an intracellular signaling pathway that precedes AChR clustering and includes the tyrosine phosphorylation of AChRs.

Mercury decreases the frequency of induced but not spontaneous clustering of acetylcholine receptors.

Studies with environmental levels of various metals typically focus on observable neurological symptoms in newborns and adults. Use of the C2C12 skeletal muscle cell line as a developmental model enabled us to test whether environmental insults prevented myotube formation or the assembly of the postsynaptic component of the neuromuscular synapse. Specifically, we asked whether the inorganic metal mercury interfered with the fusion of myoblasts into myotubes, acetylcholine receptor (AChR) clustering, or the agrin signaling events that precede AChR clustering.

The pesticide methoxychlor disrupts the fusion of myoblasts into myotubes in skeletal muscle cell culture.

We studied the effect of the estrogenic pesticide methoxychlor (MXC) on skeletal muscle development using C2C12 muscle cell culture. Various concentrations of MXC or beta-estradiol (E) were added to the culture media. MXC (100 microM) disrupted myoblast fusion into myotubes, but 10 microM MXC or 10 microM E had no effect.