No abstract

A putative choline transporter (CHOT1) has been cloned from rat brain and is reported to express a high-affinity, sodium-dependent, hemicholinium-3-insensitive choline transporter in oocytes. A second transporter (OCCREATRA) cloned from rabbit brain is 98% homologous to CHOT1 and is reported to transport creatine. We examined the distribution of CHOT1 mRNA in rat brain by in situ hybridization, using a 48 base oligonucleotide probe. In adult rats, the hybridization signal was widespread, but with a distinct pattern. High levels of expression were detected in the cerebellum (Purkinje and granule cell layers), choroid plexus, medial habenula, pontine nuclei, several brainstem nuclei, and hippocampus (pyramidal cell layer). Moderate signal was detected in cortex, globus pallidus, corpus callosum, and most other white matter tracts. Very low levels were present in striatum, nucleus accumbens, hippocampus molecular layer, and cerebellar molecular layer. Emulsion autoradiography indicated cellular localization to both neurons and glia. CHOT1 mRNA was relatively abundant in some cholinergic regions, including the medial habenula, the medial septum, and several brainstem nuclei. However, the overall pattern was distinctly different from that expected for cholinergic markers and correlated well with the localization of creatine kinase. The widespread distribution and poor correlation with cholinergic markers indicates that the CHOT1 gene does not encode the classical choline transporter known to be associated with acetylcholine synthesis. It is possible that CHOT1 is associated with cholinergic neurotransmission in some brain regions. However, it appears to encode that the rat creatine transporter, and its widespread and heterogeneous distribution suggests regions where creatine phosphate is an important energy source.

The present study examined the question of whether modulation of estradiol-induced LH surges by progesterone or antiestrogens in the immature rat might be related to changes in the concentration of pituitary GnRH receptors (GnRH-R). Rats (28 days old) that received estradiol implants at 0900 h had LH surges approximately 32 h later. Administration of progesterone or nafoxidine (U-11,100 A; 1-(2-[P-(3,4-dihydro-6-methoxy-2-phenyl-1-naphthyl)phenoxy]pyrrolidine hydrochloride) concomitantly with estradiol led to blockade of these LH surges (progesterone or nafoxidine inhibition), while progesterone treatment 24 h after estradiol brought about premature and enhanced LH release (progesterone facilitation). GnRH-R-binding capacity was determined by saturation analysis in homogenates of single pituitaries from immature rats treated with estradiol and progesterone or nafoxidine and controls treated only with estradiol using [125I]iodo-(D-Ala6,Des-Gly10)GnRh ethylamide. The affinity of GnRH-R for this analog ranged from 8.2-15.1 X 10(9) M-1 and was not affected by in vivo steroid or antiestrogen treatment. The number of GnRH-R in gonadotrophs from untreated 28-day-old rats (57.2 +/- 2.6 fmol/pituitary or 177 +/- 11 fmol/mg protein) was comparable to values previously reported for 30 day-old females. GnRH-R levels were first measured 1, 8, 24, 32, and 48 h after estradiol treatment. The pituitary content of GnRH-R paralleled changes in total pituitary protein (nadir at 24 h, rebound at 32 h, continued increase at 48 h), while their concentration (femtomoles per mg protein) was highest at 8 h. Next, GnRH-R levels were examined at 1200 h and at hourly intervals (1400-1800 h) on the afternoon of the LH surge. While GnRH-R concentrations were significantly lower at 1400 and 1700 h than at 1200 or 1800 h in animals treated with estradiol in the progesterone facilitation model, they did not change over time in the other two paradigms. There was no significant difference in pituitary content or concentration of GnRH-R at any time between immature rats treated with estradiol and progesterone or nafoxidine and their respective estradiol-treated controls. These results suggest that changes in GnRH-R levels in pituitary gonadotrophs do not play a major role in enhancement of LH surges by progesterone or in their suppression by progesterone or nafoxidine in the immature rat; therefore, these compounds may affect the pituitary at a site distal to the GnRH receptor.