External imaging of cerebral muscarinic acetylcholine receptors.

A radioiodinated ligand that binds to muscarinic acetylcholine receptors was shown to distribute in the brain by a receptor-mediated process. With single-photon-emission imaging techniques, radioactivity was detected in the cerebrum but not in the cerebellum, whereas with a flow-limited radiotracer, radioactivity was detected in cerebrum and cerebellum. Single-photon-emission computed tomography showed good definition of the caudate putamen and cortex in man.

Dopamine: release from the brain in vivo by amantadine.

After dopamine stores in the caudate nucleus of cats were labeled with [(3)H]dopamine, the ventricular system was perfused with artificial cerebrospinal fluid. The addition of amantadine to the perfusing fluid caused a doserelated increase in the concentrations of [(3)H]dopamine appearing in the perfusion effluent. Subthreshold concentrations of amantadine also enhanced the efflux of [(3)H]dopamine induced by electrical stimulation of the caudate nucleus.

Norepinephrine and dopamine: assay by mass fragmentography in the picomole range.

Gas chromatography-mass spectrometry makes possible the simultaneous measurement of norepinephrine and dopamine in concentrations of 0.1-milligram tissue samples. Specificity of the assay is confirmed both by the retention time of the compound and by the mass to charge ratio of the fragments recorded. The sensitivity is of the order of 0.5 picomole, and linearity of the response is maintained up to at least 200 picomoles.

Bimodal distribution of dopamine receptor densities in brains of schizophrenics.

The dopamine hypothesis of schizophrenia was examined by measuring the density of dopamine receptors in the postmortem brains of 81 control subjects and 59 schizophrenics from four different countries. The densities of dopamine receptors in the tissues from the schizophrenic patients had a bimodal distribution in the caudate nucleus, putamen, and nucleus accumbens. One mode occurred 25 percent above the control density, and a second mode occurred at a density 2.3 times that of the control density for all three regions. Although almost all the patients had been medicated with neuroleptics, the two modes had the same dissociation constant for the labeled ligand used, suggesting that the neuroleptic doses were similar for the two populations of schizophrenics. The results thus provide direct evidence for two distinct categories of schizophrenia.

Angiotensin-forming enzyme in brain tissue.

A renin-like enzyme is present in brain tissue and is independent of kidney and plasma renin. In the presence of homologous substrate it forms angiotensin. Administration of aldosterone significantly decreases this angiotensinforming enzyme activity, while administration of progesterone markedly enhances it.

( 3 H)lysergic acid diethylamide: cellular autoradiographic localization in rat brain.

Intravenous administration of [(3)H]lysergic acid diethylamide(LSD) to rats resulted in accumulation of the drug in the brain within 15 minutes. Autoradiographic methods were used to differentiate free and bound [(3)H]LSD in brain tissue. Free [(3)H]LSD was generally distributed in the pituitary and pineal glands, cerebellum, hippocampus,and choroid plexus. Bound [(3)H]LSD was localized in neurons of the cortex, caudate nucleus, midbrain, and medulla,as well as in choroid plexus epithelium.

Increased brain dopamine and dopamine receptors in schizophrenia.

In postmortem samples of caudate nucleus and nucleus accumbens from 48 schizophrenic patients, there were significant increases in both the maximum number of binding sites (Bmax) and the apparent dissociation constant (KD) for tritiated spiperone. The increase in apparent KD probably reflects the presence of residual neuroleptic drugs, but changes in Bmax for tritiated spiperone reflect genuine changes in receptor numbers. The increases in receptors were seen only in patients in whom neuroleptic medication had been maintained until the time of death, indicating that they may be entirely iatrogenic. Dopamine measurements for a larger series of schizophrenic and control cases (n greater than 60) show significantly increased concentrations in both the nucleus accumbens and caudate nucleus. The changes in dopamine were not obviously related to neuroleptic medication and, unlike the receptor changes, were most severe in younger patients.

Chromogranin A biosynthetic cell populations in bovine endocrine and neuronal tissues: detection by in situ hybridization histochemistry.

Chromogranin A is a highly acidic protein that is found in the secretory granules of many endocrine and neuronal cells. To localize bovine cell populations involved in chromogranin A biosynthesis, the distribution of the mRNA encoding this protein was determined with in situ hybridization histochemistry. In the adrenal gland, the mRNA was found in the chromaffin cells of the medulla but was absent from the cortex. The distribution of the mRNA in the medulla was uneven; cells located at the periphery were more heavily labeled than those in the center of the gland. Because the adrenal medulla is composed of several cell types, the chromogranin A-containing cells were further characterized for the presence of neuropeptide and adrenergic markers. Adjacent sections were examined for the mRNAs encoding enkephalin and phenylethanolamine N-methyltransferase (PNMT), the enzyme that catalyzes the formation of epinephrine from norepinephrine. Both mRNAs were present in a narrow band of cells at the periphery of the medulla. However, in contrast to the distribution of chromogranin A mRNA, the enkephalin and PNMT mRNAs were detected in only a small number of cells in the inner medullary region. The difference in the distribution of the enkephalin and PNMT mRNAs from that of chromogranin A suggests that the expression of these genes is differentially regulated. In addition to the adrenal gland, chromogranin A mRNA is expressed by many other tissues. In the parathyroid gland, which is rich in the mRNA but exhibits little chromogranin A-like immunoreactivity, the message was present in most cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Decreased serotonin S2 and increased dopamine D2 receptors in chronic schizophrenics.

Serotonin S2 and dopamine D2 receptors in the prefrontal cortex and caudate nucleus of postmortem brains of chronic schizophrenics were studied using 3H-ketanserin and 3H-spiperone, respectively. In the prefrontal cortex of schizophrenics, we found a significant decrease in the maximum number of 3H-ketanserin binding sites (Bmax), with no change in the dissociation constant (Kd). Conversely, both Bmax and Kd of 3H-spiperone binding to the caudate nucleus were significantly increased in the schizophrenic patients. There were no differences in receptor indices between patients who were taking neuroleptics until their death and those who had taken none for 2 months or more prior to death. These findings suggest that alterations in S2 receptors in the prefrontal cortex may reflect the disease process, per se, and that the increase in the number of D2 receptors in the caudate nucleus of schizophrenics is not due solely to neuroleptic medication.

Decreased receptor-binding sites for kainic acid in brains of patients with Huntington's disease.

The specific binding of (3H)kainic acid (KA) to high- and low-affinity receptor sites was assayed in postmortem samples from the brains of patients affected with Huntington's disease (HD) and age-matched controls. Treatment of rat brain with conditions that closely mimic the temperature gradient occurring in postmortem human brain only slightly but not significantly decreased receptor binding by 12 hr after death. In HD brains, total specific binding of (3H)KA was reduced in the caudate nucleus by 51%, putamen by 77%, and frontal cortex by 47%. Specific binding to the high-affinity site was virtually undetectable in the caudate nucleus and was reduced by 90% in the putamen from HD brains. No significant alterations in specific binding of (3H)KA were noted in the insular or temporal cortex, hippocampus, or cerebellum. Thus, losses of KA receptor binding were mainly localized to those regions of the HD brain that are most severely affected by neuronal degeneration, and the high-affinity receptor site appeared more affected.

Age-related decreases in dopamine receptors in the caudate nucleus and putamen of the rhesus monkey (Macaca mulatta).

Specific binding of the dopamine receptor ligand 3H-spiroperidol to cell membranes prepared from the caudate nuclei and putamens of 29 rhesus monkeys (M. mulatta), ranging in age from 2 to 22 years, was investigated. Receptor concentration (Bmax) decreased in the caudate nucleus and putamen with age at mean rates of 2.1 and 1.7% per year, respectively, whereas binding affinity (Kd) did not change significantly with age. The rate of decline in Bmax appeared to be more rapid before adulthood and in old age than during young adulthood and middle age. These data are compared with the results from similar studies of other animal species including human, rabbit, rat, and mouse. The rate of decline in striatal dopamine receptors is closely related to the rate of aging and maximal life span of the species. It may reflect both maturational and senescent processes modifying the behavior of animals as they age.

Divergent changes in D-1 and D-2 dopamine binding sites in human brain during aging.

The density of D-1 and D-2 dopamine receptors in human caudate nucleus and putamen, obtained postmortem, were studied throughout the adult lifespan using [3H]fluphenazine as the dopamine receptor ligand. The D-1 subtype increased progressively with age in both regions, while the D-2 subtype declined in caudate nucleus. The ratio of D-1/D-2 Bmax in both regions increased from approximately 1 at age 20 to 2 by age 75. The dopamine content in putamen declined with age and was inversely correlated with D-1 receptor density. We suggest that D-1 receptor density is up-regulated by loss of dopamine during aging. The D-2 receptor density in caudate nucleus was positively correlated with choline acetyltransferase activity, suggesting that loss of intrastriatal neurons with age may contribute to the decrease in D-2 sites. These divergent changes in dopamine receptor subtypes with age result in an altered complement of dopamine receptors in older humans and may provide a basis for selective pharmacotherapy in disorders of the basal ganglia.

Characterisation of brain D2 dopamine receptors solubilised by lysophosphatidylcholine.

Brain D2 dopamine receptors have been solubilised using lysophosphatidylcholine. The inclusion of proteinase inhibitors during solubilisation enables a preparation to be obtained containing a high proportion of solubilised D2 receptors with pharmacological properties similar to those of membrane-bound D2 receptors.

Basal forebrain efferents to the medial dorsal thalamic nucleus in the rhesus monkey.

Thalamic efferent connections of the basal forebrain (BF); medial septal nucleus (MS), vertical limb of the diagonal band (VDB), horizontal limb of the diagonal band (HDB), nucleus basalis (NB), and ventral pallidum (VP) were investigated in twelve rhesus monkeys. In five animals, injections of radioactively labeled amino acids were placed in the BF. In four animals, the injections involved different divisions of the NB, HDB, and the most ventral part of the VDB. In those four cases, labeled fibers in the medial forebrain bundle were observed traveling caudally towards the hypothalamus where some turned dorsally to enter the inferior thalamic peduncle. These fibers terminated in the ventral half of the magnocellular part of the medial dorsal thalamic nucleus (MDmc). In a fifth case, the amino acid injection involved most of the MS and the VDB. Labeled fibers traveled caudally from the injection site and entered the stria medullaris. These fibers then traveled caudally before turning ventrally to terminate in the dorsal half of MDmc. To determine which of the diverse neuronal types in the BF gives rise to these thalamic projections, in two monkeys injections of horseradish peroxidase (HRP) were placed into MDmc. Labeled neurons were observed throughout the full extent of the NB, the VDB, the MS, and part of the VP. In order to determine the extent of the cholinergic input to MDmc from the BF, one of the HRP cases was processed for the simultaneous visualization of HRP, and acetylcholinesterase (AChE), the hydrolytic enzyme for acetylcholine, and a second case was processed for simultaneous visualization of HRP, and choline acetyltransferase (ChAT), the synthetic enzyme for acetylcholine. We observed that 30-50% of the HRP-labeled neurons were putatively cholinergic. In order to determine if the NB projection to MD is a collateral of the NB projection to orbital frontal cortex, one fluorescent retrograde tracer was injected into the orbital frontal cortex and one into MD. This case showed that approximately 5% of the BF neurons that project to MDmc also project to the orbital frontal cortex. These results confirm a significant subcortical projection by which the cholinergic system of the basal forebrain may influence higher cortical functions through the thalamus.

What excitotoxin kills striatal neurons in Huntington's disease? Clues from neurochemical studies.

Amino acid analyses of both caudate nucleus and putamen obtained at autopsy from patients dying with Huntington's disease (HD), and from control subjects, showed significantly decreased mean glutamate contents in the HD patients. In addition, the mean glutamate concentration was significantly increased in the CSF of living HD patients as compared with controls. Neurochemical studies also showed that neither aspartic acid, proline, 5-oxoproline, nor homocysteic acid is likely to act as a causative excitotoxin in HD. Excessive striatal glycine content, or deficient glutathione content, is unlikely to contribute to the effects of a causative excitotoxin in HD. We suggest that glutamic acid may be the proximate causative neurotoxin in the striatum in HD, as a result of an unexplained failure in the reuptake mechanism for glutamate released there as an excitatory neurotransmitter.

In situ analysis of neostriatal RNA and chromatin in Alzheimer's disease.

Scanning-integrating microdensitometry of azure B-RNA- and Feulgen-Schiff-stained tissue sections was used to measure neostriatal neuronal RNA levels and susceptibility of neuronal and oligodendrocyte chromatin to acid hydrolysis in Alzheimer's disease (AD) patients and controls. AD was associated with neuronal RNA depletion (17 to 23%) in both caudate nucleus and putamen. While neuronal chromatin was found to be more acid-labile than that of the oligodendrocytes, there were no differences in either cell type between AD and controls. These data support the existence of a macromolecular disturbance (RNA loss) occurring within neostriatal neurons, perhaps related to the extrapyramidal dysfunction of AD, but fail to demonstrate that an alteration in chromatin is responsible for this effect.

Neurogenic influence on local cerebral blood flow. Effect of catecholamines or sympathetic stimulation as correlated with the sympathetic innervation.

Local cerebral flow was measured continuously in conscious rabbits (thermoclearance technique), and PaO2 and PaCO2 were recorded by mass spectrometry. Though inhalation of CO2 increased flow in caudate nucleus and lateral geniculate body, catecholamines only had effect on caudate nucleus where isoproterenol enhanced and epinephrine and norepinephrine reduced flow. Reduction by electrical stimulation of the neck sympathetic trunk was particularly evident in the caudate. Blood flow increased markedly in both regions after preganglionic conduction blockade. The effects were correlated with a significantly lower degree of sympathetic arteriolar innervation (fluorescence histochemistry) in the lateral geniculate body compared with the caudate nucleus.

Brain amino compounds in a Huntington's disease patient on isoniazid therapy.

We describe biochemical abnormalities in autopsied brain of a patient with early Huntington's disease (HD) who died of pentobarbital overdosage while under treatment with isoniazid (INH). The brain contained hydrazine, a terminal metabolite of INH, which inhibits gamma-aminobutyric acid (GABA) aminotransferase. GABA content in the basal ganglia was higher than expected for HD, and GABA content was supranormal in some brain regions. Homocarnosine (GABA-histidine) content was greatly elevated in all brain regions, suggesting chronic GABA elevation in life. Therefore, the increase in brain GABA content observed in experimental animals given INH or hydrazine also occurs in human patients.

Distribution of neuropeptides in the limbic system of the rat: the bed nucleus of the stria terminalis, septum and preoptic area.

The distribution of the neuropeptides vasoactive intestinal polypeptide, cholecystokinin octapeptide, substance P, neurotensin, methionine-enkephalin and somatostatin has been mapped immunocytochemically in the bed nucleus of the stria terminalis, one of the major sites of termination for efferent projections from the amygdala. Immunoreactive fibres and terminals were distributed more or less topographically and largely in accordance with the previously described localization of peptide-containing cell bodies in the amygdala and the amygdaloid projection fields in the bed nucleus as described by neuroanatomical techniques. Thus, vasoactive intestinal polypeptide, which was found in some of the lateral amygdaloid nuclei, had a substantial projection to the lateral bed nucleus. The lateral bed nucleus also contained cholecystokinin-octapeptide, substance P, neurotensin and methionine-enkephalin immunoreactivity which probably derived from the central amygdaloid nucleus, whilst cholecystokinin-octapeptide, and especially substance P-containing fibres, were found in the medial bed nucleus and probably arise from cells in the medial amygdala. Reciprocal amygdalopetal projections were suggested by the presence of substance P- and somatostatin-containing cell bodies in the mediodorsal bed nucleus and vasoactive intestinal polypeptide cells in the lateral bed nucleus, but somatostatin otherwise had a widespread distribution. Numerous local peptidergic connections were also noted both within the bed nucleus and between it and adjacent structures, especially the preoptic area, hypothalamus and the basal ganglia. These data provide further evidence that neuropeptides play a major role in the connectivity of the limbic system and show that the bed nucleus of the stria terminalis is an important relay station, particularly between amygdaloid efferents and other forebrain areas.

Striatal neurons expressing somatostatin-like immunoreactivity: evidence for a peptidergic interneuronal system in the cat.

Neurons expressing immunoreactivity to antisera against somatostatin 14 and other somatostatin-related peptides were identified in the striatum of cats and nonhuman primates. In each species, immunoreactive neurons were distributed singly and in small groups in the caudate nucleus, putamen and ventral striatum. A detailed study was made of somatostatin-positive neurons and neuropil in the caudate nucleus of the cat. First, the mean diameters and surface areas of neurons expressing immunoreactivity to somatostatin 14 were made from peroxidase-antiperoxidase stained material. Second, fluorescence immunohistochemistry was combined with retrograde labeling of striatal neurons to determine whether such somatostatin 14-positive neurons emit axons projecting out of the striatum. Third, the distributions of neurons and neuropil expressing immunoreactivity to somatostatin 14 or somatostatin 28 (1-12) were plotted in relation to the locations of acetylcholinesterase-poor zones ("striosomes") visible in adjoining sections. The morphometric analysis suggested that somatostatin 14-positive neurons in the caudate nucleus form a single population of medium to medium-large neurons having mean diameters of 20 micron and mean surface areas of 154 micron2. The retrograde tracer study suggested that these somatostatin 14-positive neurons are interneurons. Injections of fast blue into all of the known targets of striatofugal fiber projections failed to label somatostatin 14-positive neurons save in a few instances (less than 0.3% of more than 4000 neurons) in each of which labeling was equivocal. Analysis of the distribution of somatostatin-positive neurons and neuropil in the striatum demonstrated that both observe striosomal ordering. Somatostatin immunoreactive neuropil was dense outside and weak inside identified striosomes, and most immunoreactive neurons lay outside. Often somatostatin-positive neurons lay beside, and sometimes striosomes partly rimmed them. The processes of such neurons tended to run along the borders of the striosomes without crossing them, but occasionally single processes and rarely entire dendritic trees crossed from one compartment to the other. These results suggest that, in the striatum of the cat, somatostatin is present: (1) in fibers organized according to the compartmental distribution already recognized for other neurochemical compounds in the striatum as well as for its afferent and efferent systems, and (2) in interneurons, mostly present in the extrastriosomal matrix, but also located near striosomes, where they could serve as interfaces between the striosomes and extrastriosomal matrix.