Abstract
We use a flow-through cell perifusion system allowing continuous sampling of Luteinizing Hormone (LH) release from excised pituitary tissue. With this system, we are able to sample from viable tissue in primary culture for three days as opposed to a more typical of primary culture of 6–12 hours. Rat pituitary tissue was perifused and challenged with a physiological dose of LHRH. Samples were collected for 3.5 hours on each of 3 consecutive days in a refrigerated fraction collector and assayed for LH concentration by radioimmunoassay. In these experiments we tested two types of media, each with or without fetal calf serum supplementation, for their ability to support pituitary tissue for extended lengths of perifusion. In addition, we broadened the use of aseptic technique, sterile media and equipment to limit microbial contamination. Our results indicate that perifusion of pituitary tissue can be done successfully for 3 or more days at a modest cost, greatly increasing the amount of information that can be collected from each piece of tissue harvested.
Similar content being viewed by others
References
Carmel PW, Araki S, Ferin M (1976). Pituitary stalk portal blood collection in rhesus monkeys: evidence for the pulsatile release of gonadotropin releasing hormone (GnRH). Endocrinology 99: 243-248.
Clarke IJ, Cummins JT (1982). The temporal relationship between gonadotropin releasing hormone (GnRH) and luteinizing hormone (LH) secretion in ovarectomized ewes. Endocrinology 116: 2376-2383.
Freshney RI (1992). Towards serum free, chemically defined media for mammalian culture. In: Rickwood D and Hanes BD (eds), Animal Cell Culture (pp 15-46). New York: Oxford University News.
Gallardo E, Ramirez VD (1977). A method for the superfusion of rat hypothalami: secretion of luteinizing hormone releasing hormone (LHRH). Proc Soc Exp Biol Med 155: 79-84.
Gerhardt P, Murray RGE, Wood WA, Krieg NR (1994). Methods for general and molecular bacteriology. Washington, DC: American Society for Microbiology Press.
Knobil E (1980). The neuroendocrine control of the menstrual cycle. Recent Prog Horm Res 36: 53-88.
Levine JE, Ramierez VD (1980). In vivo release of luteinizing hormone releasing hormone estimated with push-pull cannula from the mediobasal hypothalami of ovariectomized, steroid primed rats. Endocrinology 107: 172-178.
Levine JE, Wolfe AM, Porkka-Heiskanen T, Meredith JM, Norgle JR, Turek FW (1994). In vivo sampling and administration of hormone pulses in rodents. In: Levine JE (ed.), Pulsatility in Neuroendocrine Systems (pp 129-161). New York, NY: Acedemic Press.
Moenter SM, Caraty A, Locatelli A, Karsch FJ (1991). Pattern of gonadotropin releasing hormone (GnRH) secretion leading up to ovulation in the ewe: existence of a preovulatory GnRH surge. Endocrinology 129: 1175-1182.
Pablo FD, Scanes CG, Weintraub BD (1993). History and development of radioimmunoassay. In: Pablo (ed), Handbook of Endocrine Research Techniques (pp 4-5). San Diego, CA: Acedemic Press.
Sarker DK (1987). In vivo secretion of LHRH in ovarectomized rats is regulated by a possible autofeedback mechanism. Neuroendocrinology 45: 510-513.
Tao NK, Tang SH, Chang LW (1997). A cell perifusion system of isolated islets for studies on rapid insulin-glucose dynamics. Med Eng and Physics 19: 759-764.
Urbanski HF, Pickle RL, Ramirez VD (1988). Simultaneous measurement of gonadotropin-releasing hormone, leuteinizing hormone, and follicle stimulating hormone in the orchidectomized rat. Endocrinology 123: 413-419.
Wildt L, Hausler A, Marshall G, Hutchison JS, Plant TM, Belchetz PE, Knobil E (1981). Frequency and amplitude of gonadotropin releasing hormone stimulation and gonadotropin secretion in the rhesus monkey. Endocrinology 109: 376-385.
Woller M, Nichols E, Herdendorf T, Tutton D (1998). Release of luteinizing hormone-releasing hormone from enzymatically dispersed rat hypothalamic explants is pulsatile. Biol Reprod 59: 587-590.
Woller M, Tessmer S, Neff D, Ada Nguema A, Van Roo B, Waechter-Brulla D (2001). Leptin stimulates gonadotropin releasing hormone release from cultured intact hemihypothalami and enzymatically dispersed neurons. Exp Biol Med 226: 591-596
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Vella, S., Gussick, J., Woller, M. et al. Modification of cell perifusion for extended study of hormone release in the rat pituitary. Methods Cell Sci 23, 197–204 (2001). https://doi.org/10.1023/A:1016339932088
Issue Date:
DOI: https://doi.org/10.1023/A:1016339932088