In the present study we have examined the in vivo effects of
thyroid hormone and TRH on secretory tissue concentrations of TRH and
TRH-Gly (
pGlu-His-Pro-Gly), a TRH precursor. Within secretory granules,
TRH-Gly is converted to TRH through alpha-amidation of the C-terminal
proline residue, using Gly as the NH2 donor. Using specific RIA, we measured the
TRH-Gly immunoreactivity (
TRH-Gly-IR) and TRH-IR concentrations in tissues from the reproductive and gastrointestinal systems, adrenals, and other internal organs in euthyroid, hypothyroid, and T4-treated 250-g Sprague-Dawley male rats.
TRH-Gly-IR concentrations were more than 2-fold higher than TRH-IR concentrations within the adrenal, pancreas, bowel, and stomach at the time of death. Untreated
hypothyroidism and exogenous TRH significantly increased adrenal
TRH-Gly-IR levels. Pancreatic
TRH-Gly levels increased about 2-fold in hypothyroid rats. Incubation at 60 C significantly increased
TRH-Gly-IR levels in the pancreas, adrenal, bowel, stomach, and epididymis by 14-, 3-, 6-, 6-, and 6-fold, respectively. Also after 60 C incubation increases in the
TRH-Gly-IR/TRH-IR ratio of 2.7-, 4-, and 1.7-fold were observed in the pancreas, epididymis, and bowel, respectively. Pooled
tissue extracts were fractionated by
cation exchange and reverse phase HPLC for characterization of
TRH-Gly-IR. Both chromatographic methods revealed a major peak of
TRH-Gly-IR coeluting with synthetic
TRH-Gly. Incubation at 60 C caused 13.5-, 4.1-, 1.5-, and 5-fold increments in the
TRH-Gly-IR for adrenal, pancreas, prostate, and thyroid, respectively, compared to the immediately extracted control aliquots.
Cation exchange and reverse phase HPLC also revealed production of higher mol wt TRH precursor
peptides after incubation at 60 C for 4 or 20 h. Only the
TRH-Gly-IR peak coeluting with
pGlu-His-Pro-Gly was converted into TRH by rat brain alpha-amidating
enzyme. The data suggest that biosynthesis of TRH occurs in rat extrahypothalamic tissues and may be modulated by thyroid status, iv TRH, and selective thermal inactivation of
enzymes that convert
prepro-TRH to TRH.