Modulation of electrically evoked acetylcholine release in cultured rat septal neurones.
The electrically evoked release of acetylcholine and its modulation via auto- and heteroreceptors were studied in primary cell cultures prepared from embryonic rat septum (ED 17). Cultures were grown for 1, 2 or 3 weeks on circular, poly D-lysine-coated glass coverslips. They developed a dense network of non-neuronal and neuronal cells, only some of which were immunopositive for choline acetyltransferase. To measure acetylcholine release, the cells on the coverslips were pre-incubated with [3H]choline (0.1 micromol/L), superfused with modified Krebs-Henseleit buffer at 25 degrees C and electrically stimulated twice for 2 min (S1, S2; 3 Hz, 0.5 ms, 90-100 mA). The electrically evoked overflow of [3H] from the cells consisted of approximately 80% of authentic [3H]Ach, was largely Ca2+-dependent and tetrodotoxin sensitive, and hence represents an action potential-evoked, exocytotic release of acetylcholine. Using pairs of selective agonists and antagonist added before S2, muscarinic autoreceptors, as well as inhibitory adenosine A1- and opioid mu-receptors, could be detected, whereas delta-opioid receptors were not found. Evoked [3H] overflow from cultures grown for 1 week, although Ca2+ dependent and tetrodotoxin sensitive, was insensitive to the muscarinic agonist oxotremorine, whereas the effect of oxotremorine on cells grown for 3 weeks was even more pronounced than that in 2-week-old cultures. In conclusion, similar to observations on rat septal tissue in vivo, acetylcholine release from septal cholinergic neurones grown in vitro is inhibited via muscarinic, adenosine A1 and mu-opioid receptors. This in vitro model may prove useful in the exploration of regulatory mechanisms underlying the expression of release modulating receptors on septal cholinergic neurones.
Ehret
A
A
Institut für Pharmakologie und Toxikologie, Neuropharmakologisches Laboratorium, Freiburg, Germany.
Haaf
A
A
Jeltsch
H
H
Heimrich
B
B
Feuerstein
T J
TJ
Jackisch
R
R
D000109:Acetylcholine / Q000378:metabolism*; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002478:Cells, Cultured; D002794:Choline / Q000493:pharmacokinetics; D002795:Choline O-Acetyltransferase / Q000378:metabolism; D004558:Electric Stimulation; D007150:Immunohistochemistry; D009292:Narcotic Antagonists / Q000494:pharmacology; D009294:Narcotics / Q000494:pharmacology; D009474:Neurons / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D051381:Rats; D017208:Rats, Wistar; D011976:Receptors, Muscarinic / Q000187:drug effects / Q000378:metabolism; D017450:Receptors, Opioid, mu / Q000187:drug effects / Q000378:metabolism; D018047:Receptors, Purinergic P1 / Q000378:metabolism; D020665:Septum of Brain / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D013779:Tetrodotoxin / Q000494:pharmacology; D014316:Tritium
D009292:Narcotic Antagonists; D009294:Narcotics; D011976:Receptors, Muscarinic; D017450:Receptors, Opioid, mu; D018047:Receptors, Purinergic P1; D014316:Tritium; D013779:Tetrodotoxin; D002795:Choline O-Acetyltransferase; D002794:Choline; D000109:Acetylcholine; D002118:Calcium