Comparison of peptide release from fetal rat hypothalamic neurones cultured in defined media and serum-containing media.
Primary cultures of rat hypothalamic neurones were maintained either in a serum-supplemented medium or in a serum-free chemically defined medium for up to 6 weeks. The release of the 41 amino acid-containing peptide, corticotrophin-releasing factor (CRF-41), vasopressin (AVP) and somatostatin (SRIF) were followed using immunoassays. In response to K+ (56 mmol/l) depolarization both the quantities of peptides released and the magnitude of responses were significantly greater from cultures maintained in the fully supplemented defined medium. As a consequence, release of CRF-41 and AVP could be measured directly, without requiring the concentration step necessary for cultures grown in serum. The response to K+ depolarization increased with the age of the culture, suggesting neuronal maturation. Responses to K+ depolarization were Ca2+-dependent, and the addition of corticosterone (100 nmol/l) to the defined medium caused a significant reduction in the response of neurones secreting CRF-41 and AVP, but not those secreting SRIF, to depolarization. This suggests the retention in vitro of the responsiveness of stress-associated neuropeptides to the negative feedback effects of corticosterone. Neurones producing CRF-41 and AVP responded significantly in a dose-dependent manner to acetylcholine stimulation, whereas those producing SRIF did not. As cultures matured, the CRF-41- and AVP-producing neurones became more sensitive to acetylcholine with the maximal response at 1 nmol acetylcholine/l. In conclusion, the culture of rat hypothalamic neurones is improved in terms of peptide output when the cultures are maintained in a defined medium.(ABSTRACT TRUNCATED AT 250 WORDS)
D000818:Animals; D001127:Arginine Vasopressin / Q000378:metabolism; D001769:Blood; D002478:Cells, Cultured; D003346:Corticotropin-Releasing Hormone / Q000378:metabolism; D003470:Culture Media; D007031:Hypothalamus / Q000378:metabolism*; D009474:Neurons / Q000187:drug effects / Q000378:metabolism*; D010455:Peptides / Q000378:metabolism*; D011188:Potassium / Q000494:pharmacology; D051381:Rats; D013004:Somatostatin / Q000378:metabolism
D003470:Culture Media; D010455:Peptides; D001127:Arginine Vasopressin; D013004:Somatostatin; D003346:Corticotropin-Releasing Hormone; D011188:Potassium