Receptor autoradiography in thoracic spinal cord: correlation of neurotransmitter binding sites with sympathoadrenal neurons

Receptor autoradiography was combined with the retrograde labeling of sympathoadrenal neurons by fast blue to determine whether opiate, serotonin, catecholamine, or cholinergic binding sites could be spatially correlated with preganglionic neurons in the rat intermediolateral cell column (IML) that project to the adrenal gland. [3H]Dihydromorphine (DHM) was used for the visualization of mu opiate receptors, [3H]lysergic acid diethylamide (LSD) for serotonin receptors, [3H]para-aminoclonidine (pAC) for alpha 2-adrenergic receptors, and [3H]quinuclidinyl benzilate (QNB) for muscarinic cholinergic receptors. While qualitative assessment of autoradiograms indicated that alpha 2-adrenergic and muscarinic receptors were concentrated in the intermediate zone of the spinal cord, quantitation of grains in specific regions of the intermediate gray revealed that alpha 2-adrenergic and serotonergic receptors were more highly concentrated over sympathoadrenal preganglionic neurons than over other regions in IML or the adjacent intermediate gray matter. Information concerning the distribution of neurotransmitter-binding sites in other regions of thoracic spinal cord was also obtained. All ligands showed relatively dense binding sites in the superficial laminae of the dorsal horn, and all but [3H]DHM revealed similar densities of binding sites in the region adjacent to the central canal. Only [3H]QNB revealed a high density of binding sites in the ventral horn of the spinal cord.     

Ontogenesis of adenosine receptors in the central nervous system of the rat

The ontogeny of adenosine receptors was studied in rat brain and spinal cord using the specific ligand [3H]cyclohexyladenosine [( 3H]CHA). The [3H]CHA affinity constant (Kd) and the maximum receptor binding capacity (Bmax) were analyzed at all ages and in all CNS regions studied. Throughout development the Kd of [3H]CHA binding remained relatively stable and for cortex, cerebellum, subcortex, midbrain, brainstem and spinal cord ranged from 2.2 +/- 0.2 to 5.5 +/- 0.6 nM (mean +/- S.E.M.). In contrast, the Bmax values from 1- and 90-day animals increased by as little as 2-fold in subcortical regions and by as much as 9- and 16-fold in cortex and cerebellum, respectively. The highest density of binding sites was observed in subcortical structures and the lowest in brainstem and midbrain. In cortex, a steady increase in receptor number began at day 1 and stopped at the adult level by 21 days. In cerebellum, maximum receptor proliferation began at about 14 days and continued to adulthood. Other CNS regions showed intermediate rates of receptor development. These differences may reflect both the pattern of postnatal neurogenesis in the rat CNS and the maturation of those neural elements containing adenosine receptors.     

3H]thienyl-phencyclidine ([3H]TCP) binds to two different sites in rat brain. Localization by autoradiographic and biochemical techniques

A high affinity [3H]thienyl-phencyclidine ([3H]TCP) binding and its similarity to that of [3H]phencyclidine ([3H]PCP) have been demonstrated on whole rat brain homogenates. We now describe the regional distribution of the [3H]TCP binding sites in the rat brain with fixed sections and frozen slide-mounted sections visualized by autoradiography and with homogenates of 12 regions by direct binding experiments. The 3 techniques give a similar pattern for the [3H]TCP binding distribution and the biochemical study reveals that two distinct binding sites for [3H]TCP exist: one of high affinity (5-10 nM) in the forebrain, which should be responsible for the psychotropic effects and a second one of lower affinity (50-80 nM) in the hindbrain and the spinal cord, which should be involved in the extrapyramidal behavior induced by PCP and congeneers. Competition experiments have shown that muscarinic compounds interact only with the hindbrain receptor possibly in two different sites, although morphine interacts with a very low affinity with the forebrain's high affinity receptor. Results obtained with SKF-10,047 (N-allylnormetazocine) seem to indicate that TCP and sigma-receptors are different.     

Cellular localization of estrogen receptors on neurones in various regions of cultured rat CNS: coexistence with cholinergic and galanin receptors

Autoradiographic studies have shown that many neurones in explant cultures of rat neocortex, hippocampus, preoptic area and spinal cord express binding sites for [3H]-estradiol which are distributed over the cell bodies and primary processes. By means of immunohistochemistry, it was observed that neurones were labelled by monoclonal antibodies against estrogen alpha-receptors and a polyclonal antibody against estrogen beta-receptors. Immunoreactivity was distributed over the soma and primary processes of the cells, the nuclei being more intensely stained. Double-immunostaining revealed a colocalization of estrogen alpha- and beta-receptors on approximately half of the neurones in cultures from neocortex and hippocampus whereas in cultures from preoptic area and spinal cord only few cells were double-stained. On many neurones, a coexistence of estrogen receptors and cholinergic muscarinic or nicotinic sites was found. Furthermore, combined autoradiographic and immunohistochemical studies have shown a colocalization of receptors for estrogen and the neuropeptide [125I]-galanin. The coexistence of estrogen and cholinergic sites as well as of estrogen and galanin receptors on the same neurones are discussed with respect to neurodegenerative events such as Alzheimer's disease.     

Characterization of glucocorticoid receptors in whole and cellular subfractions of embryonic chick spinal cord

Steroid-specific binding sites for tritiated corticosterone have been localized, via autoradiography, to motoneurons in the lateral motor columns of the adult rat spinal cord. Binding sites in adult rat spinal cord have been characterized biochemically and shown to possess the properties of a putative glucocorticoid receptor. The presence of receptors for glucocorticoids in embryonic chick spinal cord was determined and their characterization undertaken as a prelude to the study of the functions under regulation by glucocorticoids during development. Assay conditions were defined and binding of [3H]dexamethasone [( 3H]Dex) to cytosols of 6- and 10-day embryonic spinal cord and cellular subfractions of 6-day spinal cord determined. Saturable, high-affinity binding of [3H]Dex to cytosols prepared from both whole 6- and 10-day spinal cords and cells of all 3 cellular subfractions of 6-day spinal cords was observed. The binding component in 10-day cytosols was (1) proteinaceous, as binding was eliminated by heating cytosols, and (2) a macromolecular species, as it displayed a sedimentation coefficient of 8S upon centrifugation in sucrose gradients. The putative receptor displayed binding specific for glucocorticoids in a competition assay, with the exception that some inhibition of binding by the androgen ligand, methyltrienolone (R1881) was observed. The binding affinity decreased as the values for KD increased from 3.4 +/- 0.9 nM on day 6 to 15.7 +/- 1.8 nM on day 10, while the values for Bmax increased from 270 to 855 fmol/mg protein over the same period.(ABSTRACT TRUNCATED AT 250 WORDS)     

Characterization and localization of adenosine receptors in rat spinal cord

Adenosine A1 receptors were characterized in membranes from rat dorsal and ventral spinal cord using [3H] cyclohexyladenosine [( 3H]CHA) and compared with those in brain. For determination of anatomical loci of adenosine A1 receptors in the dorsal and ventral spinal cord, various lesions were employed, including kainic acid injections directly into the lumbar dorsal spinal cord, spinal cord hemitransections, dorsal rhizotomies, and neonatal capsaicin treatment. In control animals a single high affinity binding component was observed in dorsal and ventral spinal cord with KD values of 2.3 and 2.6 nM and Bmax values of 170 and 123 fmol/mg of protein, respectively. In comparison, [3H]CHA binding to whole brain membranes exhibited KD and Bmax values of 2.3 nM and 301 fmol/mg of protein, respectively. The IC50 values for CHA, (-)-phenylisopropyl adenosine, adenosine-5'-ethylcarboxamide, 2-chloroadenosine, (+)-phenylisopropyl adenosine, and theophylline to displace [3H]CHA were 3.6, 2.3, 15, 17, 21, and 30,500 nM for dorsal horn and 5.1, 2.7, 9.8, 24, 25, and 21,000 nM for ventral horn. The potencies of the various ligands are similar to those found for brain tissue. Injection of kainic acid directly into the dorsal spinal cord significantly reduced specific [3H]CHA binding by 33% in this tissue when compared to values from saline-injected control animals. This decrease was accompanied histologically by the depletion of intrinsic neuronal cell bodies and extensive gliosis at the injection site. Terminals of descending or primary afferent systems appear not to contain [3H]CHA-binding sites since lesions which interrupt these systems failed to alter the levels of [3H]CHA receptors in denervated spinal cord tissue.(ABSTRACT TRUNCATED AT 250 WORDS)     

Binding of the neurotrophic peptide Org 2766 to rat spinal cord sections is affected by a sciatic nerve crush

The binding of the neurotrophic peptide, [3H]Org 2766 (55 nM), to rat spinal cord sections was studied, employing quantitative autoradiography. The binding was unevenly distributed over spinal cord structures and was displaceable by non-labelled Org 2766 to a limited extent (35%). Binding could not be displaced by the opiate antagonist, naloxone, indicating that [3H]Org 2766 binding sites are distinct from opiate receptors. However, the exact nature of the binding sites remains to be elucidated. A marked left-right difference in [3H]Org 2766 binding in the dorsal horns of the spinal cord at level L2 was observed, 6 days after unilateral crush lesioning of the sciatic nerve. No such effect was found at level T10. After 28 days, when sensorimotor functioning had completely recovered, the [3H]Org 2766 binding pattern was comparable to that in sham-operated rats again. It is suggested that Org 2766 binds to axonal sprouts or glia in the dorsal horn of the spinal cord.     

Estrogen binding and the actions of testosterone in the brain of the male rhesus monkey

Autoradiography and high performance liquid chromatography (HPLC) were used to determine where metabolites of testosterone interact with estrogen binding sites in the brain of the male primate. Three days after castration, animals received a subcutaneous injection of either estradiol benzoate (EB, 200 micrograms/kg, n = 4) or oil vehicle (controls, n = 4). Three hours later, 5 mCi [3H]testosterone was administered as an intravenous bolus. At 60 min, brains were rapidly removed, left halves were used for autoradiography and right halves were dissected into 14 samples for HPLC of nuclear and supernatant fractions. In control males, labeled neurons were observed in preoptic area, hypothalamus and amygdala. In EB-pretreated males, the number of labeled neurons was reduced by 35% in the anterior hypothalamus and ventromedial nucleus, and by 65% in the cortical and accessory basal amygdaloid nuclei, but was not significantly reduced in other brain regions. In hypothalamus, preoptic area and amygdala, EB-pretreatment reduced nuclear concentrations of [3H]estradiol to 37-55% of control levels, but reduced neither the nuclear concentrations of [3H]testosterone nor the supernatant concentrations of [3H]estradiol and [3H]testosterone. The data suggest that the actions of testosterone in regions such as the arcuate nucleus and lateral septal nucleus primarily involve unchanged testosterone or dihydrotestosterone, while in regions such as the amygdala, aromatization and interaction with estrogen receptors is involved also.     

Localization of [H]nitrobenzylthioinosine binding sites in rat spinal cord and primary afferent neurons

The distribution of [3H]nitrobenzylthioinosine ([3H]NBI) binding to nucleoside transport sites in rat spinal cord and spinal roots was examined using membrane binding and autoradiographic techniques. A single class of high affinity binding sites having dissociation constants (KD) between 0.42 +/- 0.05 and 0.088 +/- 0.012 nM was observed in dorsal and ventral spinal cord and their associated roots. The maximal number of binding sites (Bmax) in dorsal and ventral spinal cord was 110.1 +/- 7.1 and 73.6 +/- 7.5 fmol/mg protein, respectively. The highest levels of [3H]NBI binding were found in the dorsal grey matter of the cervical and lumbar enlargements. Autoradiographic studies showed that [3H]NBI sites were especially concentrated in the substantia gelatinosa of the dorsal spinal cord and the nucleus caudalis of the spinal trigeminal nucleus. The level of these binding sites in dorsal roots was nearly 4 times that observed in ventral roots; 98.5 and 23.0 fmol/mg protein, respectively. Adult animals depleted of unmyelinated sensory fibers by neonatal capsaicin treatment showed significantly reduced numbers of [3H]NBI sites (35%) in dorsal roots but not ventral roots, while KD values were unaffected. These results indicate that [3H]NBI sites are enriched in areas of the spinal cord and brainstem which subserve sensory functions and that these sites are located, in part, on unmyelinated primary afferent fibers.     

Glycine high affinity uptake and strychnine binding associated with glycine receptors in the frog central nervous system

Accumulation of [3H]glycine into synaptosomal fractions occurs by high affinity systems in cerebral cortex, optic tectum, brain stem and spinal cord of the frog. Specific [3H]strychnine binding which appears associated with postsynaptic glycine receptors is also demonstrable in these regions. By contrast, only very low levels of strychnine binding and high affinity glycine uptake occur in higher centers of the rat central nervous system. The relative potencies of small neutral amino acids in competing for [3H]strychnine binding are similar in frog brain and spinal cord. No evidence for a high affinity accumulation of [3H]taurine by synaptosomal fractions of frog spinal cord can be demonstrated. These observations favor glycine rather than taurine as an inhibitory transmitter in frog spinal cord. Moreover, these findings suggest that glycine may have a synaptic role in higher brain centers in the frog.     

Binding properties of [3H]gacyclidine (cis(pip/me)-1-[1-(2-thienyl)-2-methylcyclohexyl]piperidine) enantiomers in the rat central nervous system

Gacyclidine (cis(pip/me)-1-[1-(2-thienyl)-2-methylcyclohexyl]piperidine) is a TCP derivative, which exhibits potent neuroprotective properties against glutamate-induced neurotoxicity in vitro and in vivo. In order to better understand gacyclidine pharmacological properties, the binding parameters of its enantiomers ((-) and (+)[3H]GK11) were determined in the rat central nervous system (CNS). An autoradiographic study has shown that their binding distributions are correlated with those of N-methyl-D-aspartate (NMDA) receptors throughout the CNS. Globally, the labeling was the highest with (-)[3H]GK11. In the cerebellum, both radioligands similarly labeled the molecular layer. For both radioligands, on telencephalic, cerebellum and spinal cord homogenates, the association and dissociation kinetics were accounted for by multiphasic process. In all regions, (-)[3H]GK11 exhibited the highest affinity in the nanomolar range. The pharmacological study revealed that both enantiomers labeled both high and low affinity sites in all regions. The pharmacological profile of high affinity sites was correlated with those of NMDA receptors. Those of low affinity sites were different in telencephalic and cerebellar homogenates. Overall, this study showed that low affinity sites might constitute a heterogeneous population, which could include sigma receptors in the cerebellum. The autoradiographic study has shown that these sites may be located in the molecular layer. The contribution of low affinity sites to the neuroprotective properties of gacyclidine remains to be investigated.     

Correlation of ontogeny with function of [3H]U69593 labelled kappa opioid binding sites in the rat spinal cord

In this study, we have used a variety of in vitro and in vivo techniques to demonstrate the presence, and examine the function, of [3H]U69593 binding sites in the spinal cord of the 9-16-day-old rat in comparison to the adult. Equilibrium binding of [3H]U69593 to homogenates of adult rat spinal cord revealed a single population of non-interacting sites with a maximum binding capacity of 10.4 +/- 1.4 fmol/mg protein and an apparent equilibrium dissociation constant of 2.31 +/- 0.47 nM while in 9-16-day-old cord these parameters were 57.0 +/- 9.4 fmol/mg protein and 2.28 +/- 0.22 nM, respectively. The total binding capacity per cord was 95.8 +/- 8.3 and 121.8 +/- 7.7 fmol/cord for adult and immature rat, respectively. Competition studies using receptor-selective opioid ligands showed that these sites were kappa opioid in nature. Autoradiographical techniques demonstrated a uniform distribution of these sites over transverse sections of 9-16-day-old rat cord. In vitro electrophysiology was performed on spinal cord slice preparations from the 9-16-day-old rat. U69593 (100 nM-1 microM) had no effect on passive membrane properties but produced a naloxone-reversible depression of both spontaneous and electrically evoked activity in dorsal horn neurones. Direct intrathecal injection of U69593 (0.3-10.0 micrograms/animal) into 9-16-day-old rats produced a dose-dependent, naloxone-reversible, antinociception when measured using the paw-pressure test. In conclusion, we have shown that, in contrast to the adult, the spinal cord of the 9-16-day-old rat has a significantly higher concentration of [3H]U69593 binding sites which have functional in vitro and in vivo correlates.     

Biochemical characterization and regional quantification of mu, delta and kappa opioid binding sites in rat spinal cord.

The spinal cord contains mu, delta and kappa opioid receptors which mediate the antinociceptive effects of opioid agonists administered onto the spinal cord. In this study, we characterized the binding sites for highly-selective mu, delta and kappa opioid radioligands and quantified the distribution of opioid binding sites in rat lumbosacral spinal cord using autoradiography. In sections of rat brain mounted on glass slides, the mu ligand, [3H]sufentanil, bound with high affinity with an apparent Kd of 0.46 nM. The delta ligand, [3H]DPDPE [( D-Pen2.5]-enkephalin), bound with a Kd of 4.31 nM, and the kappa-ligand, [3H]U69593, bound with a Kd of 2.27 nM. Three regions of the spinal gray were targeted for quantification of binding sites by autoradiography. The data indicate that when considered as a percentage of the total opioid binding capacity within a region, the contribution of mu sites in laminae I-II was about 90%, with delta and kappa sites 7% and 3%, respectively. In lamina V, the mu sites comprised about 70% of the total opioid sites, with delta and kappa sites comprising 28% and 2%, respectively. In the area adjacent to the central canal, mu sites contributed about 65% of the total opioid sites followed by delta sites at 33% and kappa sites at 2% of total opioid sites. These results demonstrate a differential distribution of mu, delta and kappa binding sites with respect to the organization of the spinal gray matter. The preferential occurrence of all 3 opioid binding sites in the superficial dorsal horn is noteworthy since many fine caliber primary afferent fibers mediating nociception establish synaptic contact in this region.     

Characterization of [3H]rauwolscine binding to alpha 2-adrenoceptor sites in the lumbar spinal cord of the cat: comparison to such binding sites in the cat frontal cerebral cortex

The binding of the selective alpha 2-adrenoceptor antagonist radioligand [3H]rauwolscine ([3H]RAUW) to homogenates of cat frontal cerebral cortex and cat lumbar spinal cord was investigated. Experiments were performed at 20 degrees C in 50 mM Tris HCl/l mM Na2EDTA buffer (pH 6.9 at 20 degrees C). At this temperature, specific [3H]RAUW binding, defined as the difference between the amount of [3H]RAUW bound in the absence and presence of 1 microM rauwolscine or 1 microM rauwolscine or 1 microM yohimbine, reaches equilibrium values by approximately 60 min and is reversible with a mean t1/2 of dissociation of 15 min in cortex and 20 min in spinal cord. The kinetically determined Kd of [3H]RAUW (mean K-1/mean K1) was 0.59 nM and 1.68 nM in cortex and spinal cord, respectively. The results of equilibrium saturation experiments, routinely performed at [3H]RAUW concentrations between 0.1 nM and 6.0 nM, indicate that [3H]RAUW binds to saturable sites in both CNS regions of the cat. Scatchard plots of saturation isotherm data were consistently linear and the mean Kd value determined from 10 such experiments was 0.72 nM in frontal cortex and 0.82 nM in lumbar spinal cord. A mean Bmax value of 230 fmol/mg protein was determined for saturable [3H]RAUW binding sites in the cat frontal cortex. In teh cat lumbar spinal gray, a mean Bmax value for saturable [3H]RAUW binding sites of 75 fmol/mg protein was obtained. Saturable [3H]RAUW binding sites in the cat lumbar spinal gray are present at apparently equal density in dorsal and ventral horns. Inhibition experiments, performed at 0.2 nM or 0.4 nM [3H]RAUW, indicate that the binding sites labeled by [3H] RAUW possess a pharmacology characteristic of alpha-adrenoceptors. Thus, rauwolscine, yohimbine, and phentolamine compete for specific [3H]RAUW binding with high affinity and are much more potent inhibitors than corynanthine, prazosin, and propranolol. Mean Hill coefficients, calculated from logit-log plots of competition data, were close to one for all antagonists examined. L-Epinephrine and L-norepinephrine were 15-20 times more potent inhibitors of specific [3H]RAUW binding than were their corresponding D-isomers. The agonist inhibitor potency series: p-aminoclonidine = clonidine = L-epinephrine greater than L-norepinephrine much greater than isoproterenol, is that expected of alpha 2-adrenoceptor sites. Mean Hill coefficients efficients for all agonists were considerably less than one.(ABSTRACT TRUNCATED AT 400 WORDS)     

Binding properties of [H]gacyclidine (cis(pip/me)-1-[1-(2-thienyl)-2-methylcyclohexyl]piperidine) enantiomers in the rat central nervous system

Gacyclidine (cis(pip/me)-1-[1-(2-thienyl)-2-methylcyclohexyl]piperidine) is a TCP derivative, which exhibits potent neuroprotective properties against glutamate-induced neurotoxicity in vitro and in vivo. In order to better understand gacyclidine pharmacological properties, the binding parameters of its enantiomers ((−) and (+)[³H]GK11) were determined in the rat central nervous system (CNS). An autoradiographic study has shown that their binding distributions are correlated with those of N-methyl- -aspartate (NMDA) receptors throughout the CNS. Globally, the labeling was the highest with (−)[³H]GK11. In the cerebellum, both radioligands similarly labeled the molecular layer. For both radioligands, on telencephalic, cerebellum and spinal cord homogenates, the association and dissociation kinetics were accounted for by multiphasic process. In all regions, (−)[³H]GK11 exhibited the highest affinity in the nanomolar range. The pharmacological study revealed that both enantiomers labeled both high and low affinity sites in all regions. The pharmacological profile of high affinity sites was correlated with those of NMDA receptors. Those of low affinity sites were different in telencephalic and cerebellar homogenates. Overall, this study showed that low affinity sites might constitute a heterogeneous population, which could include σ receptors in the cerebellum. The autoradiographic study has shown that these sites may be located in the molecular layer. The contribution of low affinity sites to the neuroprotective properties of gacyclidine remains to be investigated.     

Sites of aromatization of [H]testosterone in forebrain of male, female and androgen receptor-deficient Tfm mice: an autoradiographic study

The distribution of radioactivity after injection of [3H]testosterone was studied in the forebrain of adult mice by thaw-mount autoradiography. Nuclear labeling was high in neurons in the dorsal part of the medial nucleus of the amygdala and in the dorsocaudal part of the bed nucleus of the stria terminalis. Low nuclear uptake occurred in the medial preoptic nucleus, in mediobasal hypothalamic nuclei and in the ventromedial amygdala. Nuclear concentration of radioactivity was not influenced by competition with dihydrotestosterone and was present in androgen receptor deficient Tfm mice. It was totally abolished by competition with estradiol. This indicates that in the brain [3H]testosterone is converted to estrogenic metabolites which bind to estrogen receptors. Nuclear labeling after [3H]testosterone was restricted to a few of the brain nuclei, known to contain estrogen receptors indicating that aromatization occurs only in select regions. The results suggest that testosterone acts on the brain via estrogen receptors.     

The glycine receptor in the mutant mouse spastic (spa): strychnine binding characteristics and pharmacology

There is a marked deficit in the binding of the glycine receptor antagonist strychnine to the CNS of the mutant mouse spastic. The characteristics and pharmacology of [3H]strychnine binding to washed homogenates of spastic and littermate control spinal cord and brainstem were investigated to determine the nature of this defect. The maximal binding of [3H]strychnine to spastic homogenates is approximately 20% the value obtained from littermate control homogenates; the affinity of [3H]strychnine binding is approximately 25% lower than littermate control values. The pharmacology of [3H]strychnine binding has the same rank order of potency in spastic and littermate control mice; however, there are small differences in the potency of several compounds in spastic vs littermate control animals. These results indicate that the glycine receptor alteration seen in the spastic mutant mouse is primarily due to a decrease in receptor number. They also suggest that the pharmacological characteristics of the glycine receptor in spastic are different from littermate control. The data do not permit a distinction between whether the strychnine binding sites found in spastic represent the same population seen in littermate control animals or are a subpopulation that is spared by the mutation.     

Distinct binding sites for calcitonin gene-related peptide and salmon calcitonin in rat central nervous system

Calcitonin gene-related peptide (CGRP) binding sites have been identified in homogenates from the rat brain and spinal cord. Autoradiography with [¹²⁵I]rat CGRP (rCGRP) revealed high grain density over the lateral hypothalamus, vestibular nuclei, colliculi, medial geniculate body, corpus mamillare and the molecular layer of the cerebellum which lacked binding sites for [¹²⁵I]salmon calcitonin (sCT). In contrast, no rCGRP labeling was seen over the anterior and dorsomedial hypothalamus which showed high sCT binding. The different regional distribution of rCGRP and sCT binding sites indicates that the structurally related peptides interact with separate receptors. The overlap between the localization of CGRP binding sites and endogenous CGRP in many regions of the central nervous system suggests that CGRP exerts unique physiological functions in the central nervous system.     

Binding of [H]glycine, [H]β-alanine and [H]strychnine in cultured rat spinal cord and brain stem

Cultures of rat brain stem and spinal cord were used to visualize binding sites for [³H]glycine, [³H]β-alanine and their antagonist [³H]strychnine by light microscopic autoradiography. In spinal cord cultures, all radio-ligands were bound mainly to large neurones, probably motoneurones whereas in brain stem cultures, both medium-sized and large neurones were labelled. In contrast, glial cells did not show binding sites for any of the compounds studied, suggesting that glial elements may not possess receptors for glycine and β-alanine.     

Autoradiographic localization of thyrotropin-releasing hormone receptors in the rat central nervous system

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.