Methionine-enkephalin content in the guinea pig myenteric plexus was determined before and after acute, short-term electrical or chemical stimulation. Stimulation at 20 Hz for 30 s or exposure to high potassium, the calcium channel agonist, CGP28 392, or the narcotic antagonist, (-)-naloxone, resulted in a significant increase in the content of myenteric methionine-enkephalin. The increase produced by electrical stimulation is dependent upon functional sodium channels and the presence of extracellular calcium. These results indicate that tissue levels of enkephalin are not fixed but can fluctuate in response to nerve stimulation and suggest a mechanism whereby the rate of production of this opioid peptide is coupled to neuronal activity. Furthermore, the ability of (-)-naloxone but not (+)-naloxone to almost double myenteric enkephalin content suggests that the neurons in which this increase occurs are under tonic modulation (direct or indirect) by opioids.