We have utilized a method to minimize cytosol progestin receptor loss during freezing in order to localize and quantify estrogen-inducible progestin receptors in individual nuclei of the female rat brain.Ovariectomized females received estradiol benzoate (20 pg for 3 days) or vehicle prior to sacrifice. All animals were perfused with cold distilled Hz0 containing the cryoprotective compound, dimethyl sulfoxide (DMSO; 10% (v/v)). Thirty-one nuclei or brain regions were removed from frozen sections (300 pm) according to the method of Palkovits (Palkovits, M. (1973) Brain Res. 59: 449-450) and were assayed in vitro using a synthetic radioligand, r3H]R5020.In ovariectomized animals perfused with DMSO, a basal level (1 to 8 fmol/mg of protein) of progestin receptors was observed in a variety of preoptic, hypothalamic, and limbic structures. Moreover, estrogen treatment induced high levels (24 to 49 fmol/mg of protein) of progestin receptors in regions of the preoptic area of hypothalamus which contain high levels of estrogen receptors. These regions included the medial, periventricular, and superchiasmatic nuclei of the preoptic area, the periventricular anterior hypothalamus, the ventromedial nucleus, and the arcuatemedian eminence. Moderate levels (2 to 8 fmol/mg of protein) of progestin receptors were induced by estrogen in other hypothalamic and limbic structures, including the anterior and lateral hypothalamus, the bed nucleus of the stria terminalis, the cingulate cortex, the medial amygdaloid nucleus, and the CA, subfield of the hippocampus. By contrast, some areas, such .as the caudateputamen and the supraoptic nucleus, were devoid of both estrogen-inducible and uninduced progestin receptors.These results support the hypothesis that progesterone action in the central nervous system is mediated by cytosol receptors in discrete brain regions and provide the first quantitative map of progestin binding in a vertebrate brain.Progesterone (P) and estradiol (E2) synergize to activate feminine reproductive behavior and to induce gonadotropin secretion in a number of vertebrate species, including the rat and guinea pig. Previous studies which have employed [3H]progesterone as a radioligand usually have failed to identify specific progestin-binding components which could mediate the central actions of P in the vertebrate brain (for references, see Feder and Marrone, 1978;McEwen, 1978).

In vitro autoradiography was used to characterize and quantitatively map the sodium-independent binding of [3H]glutamate in rat brain. Measured in the presence of chloride, glutamate binding to frozen brain sections was specific, saturable, and reversible, with a Kd in the low micromolar range. At least two distinct binding sites were detected which had different affinities for quisqualic acid (0.7 PM and 1.2 mM). Autoradiograms revealed very high levels of binding in rostra1 forebrain areas, especially olfactory structures and frontal cortex.

We employed quantitative autoradiography to examine the distribution of thyrotropin-releasing hormone (TRH) receptors in the rat CNS. The binding of [3H]3-methyl-histidine-TRH [( 3H]MeTRH) to TRH receptors in frozen rat brain sections was saturable, of a high affinity (Kd = 5 nM), and specific for TRH analogs. Autoradiograms of [3H]MeTRH binding showed highest concentrations of TRH receptors in the rhinencephalon, including accessory olfactory bulb, nuclei of the amygdala, and the ventral dentate gyrus and subiculum of the hippocampus. Moderate TRH receptor concentrations were found within the thalamus and hypothalamus, in most regions of the rhombencephalon, such as the cranial nerve nuclei, and in the substantia gelatinosa of the spinal cord. Neocortex and basal ganglia contained low densities of TRH receptors. This distribution correlates well with the sensitivity of brain regions to the known effects of TRH, and suggests that TRH receptors may mediate the actions of TRH in the rat CNS.

Little information is currently available on the localization of noradrenergic systems in the human CNS. We used quantitative autoradiography with [125I] iodopindolol to examine beta-adrenergic receptors in postmortem human brain. The concentration of beta-receptors was highest in all subfields of the hippocampus, followed by cerebellum, and then thalamic nuclei, basal ganglia, midbrain, and cerebral cortex. Low levels were found in white matter and hypothalamus. This distribution differed from the distribution of beta-receptors reported in membrane homogenates of human brain and also from the distribution of beta-receptors in rat brain determined by autoradiography. The similarities and differences between the distribution of beta-receptors in the human and rat brains may have implications regarding the role of norepinephrine in the CNS of these two species.

The amount of cytoplasmic receptor for the steroid hormone, estradiol (E2), was determined in 46 nuclei and subdivisions of rat brain. Individual nuclei were removed from 300-micrometers frozen sections according to the punch-out method of Palkovits (Palkovits, M. (1973) Brain Res. 59: 449-450), and the content of E2 receptor was measured with a sensitive radioligand binding method. Cytoplasmic receptors for E2 were distributed heterogeneously throughout the rat brain. The highest level of receptor (40 fmol/mg of protein) was found in the periventricular nucleus of the preoptic area, while low (1 fmol/mg) but detectable levels of receptors were found in such limbic regions as the nucleus of the diagonal band, the olfactory tubercle, and the cingulate cortex. Regions that were devoid of detectable receptor included the medial septum, the parietal cortex, and the ventral thalamus. Our results support the notion that E2 influences reproductive behavior and neuroendocrine function by binding to receptors in discrete areas of the brain and provide the first quantitative map of E2 receptors in individual rat brain nuclei.