Determination of Hypothalamic Hormones

Many tests substances change the secretion of hypothalamic hormones, either by direct action or more generally by a feedback effects at the hypothalamic level. Under these conditions, the tissue concentration in hypothalamic specimens of treated rats is of interest, especially because several hypothalamic peptide assays are available and can be measured in the same specimen. In contrast to the tissue concentration, for many hypothalamic hormones it is difficult to measure the circulating concentrations, due to analytical problems of low concentration and rapid inactivation by enzymes. Circulating concentrations would also change immediately due to the interference of anesthesia and of the autopsy procedure. It is therefore recommended to take samples of hypothalamic tissue, and control samples of cerebral cortical tissue. 

Specific dissection is performed on the brain and pituitary gland. Hypothalamic fragments may be dissected and shock frozen immediately in liquid nitrogen, for later extraction and determination of hypothalamic peptides. The procedure is suitable for the rat and for the rabbit. Immediately after sacrifice, the skull is opened, the brain is removed and the hypothalamic 

The pituitary gland is then dissected out and stored separately. There are numerous publications which refer to the measurement of luteinising hormone releasing hormone (LHRH - Adams and Spies 1981a-c, Aubert et al. 1980, Berault et al. 1983, Catt et al. 1980, Chan et al. 1981, Clayton 1982a,b, Clayton and Catt 

The peptide concentration in the individual specimens is determined by radioimmunoassay, using a synthetic reference peptide for the standard curve. Several commercial assays are now available for hypothalamic peptides. An alternative method is the determination of peptide content in tissue extracts by an HPLC method or by mass spectrometry. 

Group means are calculated and appropriate tests for significance are applied. 

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In repeated dose studies, hypothalamic tissue is readily available at autopsy of rats and can be used for the determination of hypothalamic hormone contents, it is important to take samples are rapidly and stop enzyme... 

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