Modulation of electrically evoked serotonin release in cultured rat raphe neurons.
Electrically evoked release of serotonin (5-HT) and its modulation via 5-HT autoreceptors and alpha(2)-heteroreceptors was studied in primary cell cultures prepared from the embryonic (ED 15) rat mesencephalic brain region comprising the raphe nuclei. Cultures were grown for up to 3 weeks on circular glass coverslips. They developed a dense network of non-neuronal and neuronal cells, some of which were positive for tryptophan hydroxylase. To measure 5-HT release, the cultures were pre-incubated with [(3)H]5-HT (in the presence of the selective noradrenaline reuptake inhibitor oxaprotiline [1 micromol/L]), superfused with modified Krebs-Henseleit medium containing 6-nitroqipazine [1 micromol/L] and electrically stimulated using two conditions. Condition A: 360 pulses, 3 Hz, 0.5 ms, 90 mA, or condition B: 4 pulses 100 Hz, 0.5 ms, 90 mA (a condition which diminishes interactions with endogenously released transmitters during ongoing stimulation). After only 1 week in culture, the electrically evoked overflow of [(3)H] was Ca(2+) dependent and tetrodotoxin sensitive, suggesting an action-potential-induced exocytotic release of 5-HT. Using stimulation condition A in cultures grown for 2 weeks, both basal and evoked 5-HT release were strongly enhanced by methiotepine (1 micromol/L) but unaffected by the 5-HT(1B) autoreceptor agonist CP-93, 129 (1 micromol/L) and the alpha(2)-adrenoceptor agonist UK-14, 304 (1 micromol/L). Conversely, using stimulation condition B, not only CP-93, 129 (IC(50) 8.1 +/- 1.4 nmol/L) and UK-14, 304 (IC(50) 14.9 +/- 1.6 nmol/L) had inhibitory effects on cells grown for 2 weeks, but also the 5-HT(1A) agonist 8-hydroxy-2-(di-n-propylamino)tetralin. In conclusion, we describe for the first time electrically evoked release of 5-HT from primary cultures of fetal raphe cells and its modulation via 5-HT(1B) and 5-HT(1A) auto- and alpha(2)-heteroreceptors. Such cultured raphe cells may represent a suitable model to study expression and development of presynaptic receptors on serotonergic neurons in-vitro.
Birthelmer
Anja
A
Institute for Experimental and Clinical Pharmacology and Toxicology, University of Freiburg, Laboratory of Neuropharmacology, Freiburg, Germany.
Ehret
Andreas
A
Riegert
Céline
C
Rothmaier
Anna Katharina
AK
Leemhuis
Jost
J
Jackisch
Rolf
R
D000818:Animals; D002118:Calcium / Q000378:metabolism; D002478:Cells, Cultured; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D004558:Electric Stimulation / Q000379:methods; D004622:Embryo, Mammalian; D005260:Female; D006897:Hydroxyindoleacetic Acid / Q000378:metabolism; D007150:Immunohistochemistry / Q000379:methods; D008376:Maprotiline / Q000031:analogs & derivatives / Q000494:pharmacology; D009419:Nerve Tissue Proteins / Q000378:metabolism; D009474:Neurons / Q000187:drug effects / Q000378:metabolism* / Q000528:radiation effects; D011247:Pregnancy; D011903:Raphe Nuclei / Q000166:cytology*; D051381:Rats; D012701:Serotonin / Q000378:metabolism*; D017367:Selective Serotonin Reuptake Inhibitors / Q000494:pharmacology; D013997:Time Factors; D014316:Tritium / Q000378:metabolism; D014365:Tryptophan Hydroxylase / Q000378:metabolism
D002118:Calcium; D006897:Hydroxyindoleacetic Acid; D008376:Maprotiline; D009419:Nerve Tissue Proteins; D012701:Serotonin; D017367:Selective Serotonin Reuptake Inhibitors; D014316:Tritium; D014365:Tryptophan Hydroxylase; C027299:hydroxymaprotilin