Corpus callosum in sexually dimorphic and nondimorphic primates.

The midsagittal area and other morphological measures were taken on the corpus callosum of four different species of primate: Macaca mulatta, M. fascicularis, Callithrix jacchus, and Saguinus oedipus. The first two species are strongly dimorphic, whereas the New World forms show little dimorphism with regard to overall body size, canines, and brain weight. Neither total corpus callosal area (TOTALCC), or other parts of the corpus callosum (CC) showed any significant sexual dimorphism in any of the primate species sampled. Only in M. mulatta did a sexual dimorphism appear to be significant. In males of this species, the dorsoventral width of the splenium was larger than in females. In addition, the anterior commissure (ANTCOMM) evinced no sexual dimorphism in the different species. Brain weight was significantly dimorphic in only M. mulatta, and when ratio data were used to correct for brain weight, no significant differences were found in the corpus callosum. This is in contrast to Homo sapiens, where the relative size of the CC has been reported to be larger in females, and particularly so in the posterior, or splenial portion of the CC. Correlation coefficients were calculated for the various variables within each species. In general, most of the callosal measures are significantly inter-correlated, although the exact pattern varies for each species. Thus, unlike Homo sapiens, or pongids such as Gorilla and Pan, neither New nor Old World monkeys show any striking evidence for sexual dimorphism in the corpus callosum.     

Relationship of squamosal suture to asterion in pongids (Pan): relevance to early hominid brain evolution.

Based on 244 measurements of the relationship of the squamosal suture to the landmark asterion in 49 chimpanzee skulls, it is shown that in the normal lateral view the squamosal suture is very rarely inferior to asterion. In hominid crania, the squamosal suture is always well superior to asterion. Even in Pan, that part of the squamosal suture most homologous with the remnant found on the Hadar AL 162-28 Australopithecus afarensis hominid cranial fragment is very rarely inferior to asterion. Such variability suggests that Falk's (Nature 313:45-47, 1985) orientation of the Hadar specimen is incorrect; she places asterion superior to the position of the squamosal suture if projected endocranially. The implication for the brain endocast is that, however the fragment is oriented, the posterior aspect of the intraparietal (IP) sulcus is in a very posterior position relative to any chimpanzee brain. The distance from the posterior aspect of IP to occipital pole is twice as great in chimpanzee brain casts than on the Hadar AL 162-28 endocast, even though the chimpanzee brain casts are smaller in overall size. This suggests that brain reorganization, at least as exemplified as a reduction in primary visual striate cortex (area 17 of Brodmann), occurred early in hominid evolution, prior to any major brain expansion.     

Cannabinoid Receptor-Regulated Cyclic AMP Accumulation in the Rat Striatum

The present study demonstrates that desacetyllevonantradol, a synthetic cannabinoid analog, reduces cyclic AMP levels in rat striatal slices stimulated with vasoactive intestinal peptide or SKF 38393, a D1-dopamine agonist. Desacetyllevonantradol and the D2 agonist LY 171555 both inhibited D1-stimulated cyclic AMP accumulation in the striatum. Spiperone, a specific D2-dopamine antagonist, fully reversed the inhibitory effect of LY 171555 but not that of desacetyllevonantradol, indicating that this cannabinoid response is not occurring through a D2-dopaminergic mechanism. Morphine also inhibited cyclic AMP accumulation in striatal slices stimulated with either SKF 38393 or vasoactive intestinal peptide. Naloxone, an opioid antagonist, fully reversed the effect of morphine but not that of desacetyllevonantradol, indicating that cannabinoid drugs are not acting via a mechanism involving opioid receptors. The response to maximally effective concentrations of desacetyllevonantradol was not additive to that of maximally effective concentrations of either morphine or LY 171555, suggesting that dopaminergic, opioid, and cannabinoid receptors may be present on the same populations of cells.     

Characterization of Neuronal and Endothelial Forms of Angiotensin Converting Enzyme in Pig Brain

The molecular forms of angiotensin converting enzyme (ACE; EC 3.4.15.1) in preparations of pig brain cortical microvessels and striatal synaptosomal membranes have been identified by immunoelectrophoretic blot analysis. The cortical microvessels contained only the endothelial form of the enzyme, Mr 180,000, which comigrated with pig kidney ACE on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. In contrast, the synaptosomal membranes contained only a smaller form of ACE, Mr 170,000, which represents the neuronal form of the enzyme. No significant differences in inhibitor sensitivity or substrate specificity were detected between the two forms of ACE. In particular, neurokinin A was resistant to hydrolysis by either microvessel or synaptosomal membrane ACE, and the pattern of hydrolysis of substance P by the two preparations was identical.     

Developmental Changes in Distribution of Acidic Fibroblast Growth Factor in Rat Brain Evaluated by a Sensitive Two-Site Enzyme Immunoassay

We developed a sensitive two-site enzyme immunoassay (EIA) system for acidic fibroblast growth factor (aFGF), using a polyclonal antibody raised in rats. This assay is based on the sandwiching of the antigen between anti-aFGF antibody immunoglobulin G (IgG) coated on plates and biotinylated anti-aFGF antibody IgG; the detection of biotinylated IgG was performed by enzyme reaction of streptavidin-conjugated beta-D-galactosidase (beta-D-galactoside hydrolase; EC 3.2.1.23). Our system was specific for aFGF, because basic fibroblast growth factor, which shares a 55% homology of amino acid sequence with aFGF, hardly cross-reacted at all. The sensitivity of this system (0.2 ng/ml) enabled us to quantify endogenous immunoreactive aFGF in the CNS. Using this two-site EIA system, we examined the levels of aFGF in various regions of rat brain and their developmental changes. At the early stage of neonatal development, i.e., 2 days after birth, all brain regions registered low aFGF levels (less than 10 ng/g tissue). However, at the young adult stage (21- to 49-day-old animals), an extremely high level of aFGF (75-90 ng/g tissue) was found in the ponsmedulla; relatively high levels (30-40 ng/g tissue) were found in the diencephalon and mesencephalon; and comparatively low aFGF levels (5-15 ng/g tissue) were found in various other brain regions such as the frontal cortex, piriform cortex, hippocampus, olfactory bulb, cerebellum, and striatum. This marked change in the regional distribution of aFGF in the rat brain during postnatal development from 2 to 21 days after birth suggests that this factor plays a significant role in the brain during this period.     

Cerebral Polyamine Metabolism in Reversible Hypoglycemia of Rat: Relationship to Energy Metabolites and Calcium

Thirty minutes of insulin-induced reversible hypoglycemic coma (defined in terms of cessation of EEG activity) was produced in anesthetized rats. At the end of the hypoglycemic coma or after recovery for 3, 24, or 72 h induced by glucose infusion, the animals were reanesthetized and their brains frozen in situ. Two control groups were used: untreated controls without prior manipulations, and insulin controls, which received injections of insulin followed by glucose infusion to maintain blood glucose within the physiological range. The brains of these latter animals were frozen 3, 24, or 72 h after glucose infusion. Tissue samples from the cortex, striatum, hippocampus, and thalamus were taken to measure ornithine decarboxylase (ODC) activity, and putrescine and spermidine levels, as well as phosphocreatine (PCr), ATP, glucose, and lactate content. In addition, 20-microns thick coronal sections taken from the striatum and dorsal hippocampus were used for histological evaluation of cell damage and also stained for calcium. Insulin in the absence of hypoglycemia produced a significant increase in ODC activity and putrescine level but had no effect on the profiles of energy metabolites or spermidine. During hypoglycemic coma, brain PCr, ATP, glucose, and lactate levels were sharply reduced, as expected. Energy metabolites normalized after 3 h of recovery. In the striatum, significant secondary decreases in PCr and ATP contents and rises in glucose and lactate levels were observed after 24 h of recovery. ODC activity, and putrescine and spermidine levels were unchanged during hypoglycemic coma. After 3 h of recovery, ODC activity increased markedly throughout the brain, except in the striatum. After 24 h of recovery, ODC activity decreased and approached control values 2 days later. Putrescine levels increased significantly throughout the brain after reversible hypoglycemic coma, the highest values observed after 24 h of recovery (p less than or equal to 0.001, compared with controls). After 72 h of recovery, putrescine levels decreased, but still significantly exceeded control values. Reversible hypoglycemic coma did not produce significant changes in regional spermidine levels except in the striatum, where an approximately 30% increase was observed after 3 and 72 h of recovery (p less than or equal to 0.01 and p less than or equal to 0.05, respectively). Twenty-four hours after hypoglycemic coma, intense calcium staining was apparent in layer III of the cerebral cortex, the lateral striatum, and the crest of the dentate gyrus. After 72 h of recovery, the intense calcium staining included also cortical layer II, the septal nuclei, the subiculum, and the hippocampal CA1-subfield.(ABSTRACT TRUNCATED AT 400 WORDS)     

Phosphoglycerates and Protein Phosphorylation: Identification of a Protein Substrate as Glucose-1,6-Bisphosphate Synthetase

We have previously reported the occurrence of two endogenous protein phosphorylation systems in mammalian brain that are enhanced in the presence of 3-phosphoglycerate (3PG) and ATP. We present here a study of one of these systems, the phosphorylation of the 72-kDa protein (3PG-PP72). This system was separated into the substrate, 3PG-PP72, and a kinase by ammonium sulfate fractionation, hydroxyapatite chromatography, and hydrophobic interaction HPLC. The substrate protein was shown to be directly phosphorylated with [1-32P]1,3-bisphosphoglycerate [( 1-32P]1,3BPG) with an apparent Km of 1.1 nM. Nonradioactive 1,3BPG inhibited 32P incorporation in the presence of [gamma-32P]ATP and 3PG. Phosphopeptide mapping and phosphoamino acid analyses indicated that the site of phosphorylation of 3PG-PP72 observed in the presence of 3PG and ATP is a serine residue identical to that observed with [1-32P]1,3BPG. Moreover, [32P]phosphate incorporated into 3PG-PP72 in the presence of 3PG and ATP was removed by subsequent incubation with glucose-1-phosphate or glucose-6-phosphate. Finally, 3PG-PP72 showed chromatographic behaviors identical to those of glucose-1,6-bisphosphate (G1,6P2) synthetase. Based upon these observations, we conclude that 3PG-PP72 is G1,6P2 synthetase and that it is phosphorylated directly by 1,3BPG, which is formed from 3PG and ATP by 3PG kinase present in a crude 3PG-PP72 preparation.     

Electroconvulsive Shock Increases α1b-but Not α1a-Adrenoceptor Binding Sites in Rat Cerebral Cortex

Repeated administration of electroconvulsive shock (ECS) increases [3H]prazosin binding to alpha 1-adrenoceptors in rat cerebral cortex. In contrast, [3H]WB4101 binding in cortex has been reported to be unchanged after ECS. [3H]Prazosin labels two alpha 1-adrenoceptor subtypes, termed alpha 1a and alpha 1b, whereas [3H]WB4101 labels the alpha 1a subtype preferentially. The purpose of this study was to determine whether ECS increases one or both alpha 1-adrenoceptor subtypes in rat cerebral cortex. We found that treatment of rats with ECS once daily for 10-12 days increased [3H]prazosin binding in cortex by about 25% but did not significantly alter [3H]WB4101 binding to alpha 1-adrenoceptors. Measurement of alpha 1a and alpha 1b receptors by competition analysis of the selective alpha 1a antagonist 5-methylurapidil against [3H]prazosin and measurement of [3H]prazosin binding in homogenates preincubated with chlorethylclonidine, which alkylates alpha 1b binding sites, also indicated that the ECS-induced increase in alpha 1-adrenoceptors is confined to the alpha 1b subtype. In contrast to its effect on [3H]prazosin binding, ECS did not increase phosphoinositide hydrolysis as measured by [3H]inositol 1-phosphate accumulation in slices of rat cerebral cortex stimulated by either norepinephrine or phenylephrine. The failure of ECS to increase [3H]inositol 1-phosphate accumulation stimulated by phenylephrine, which is a partial agonist for this response, suggests that spare receptors do not account for the apparent absence of effect of ECS on alpha 1-adrenoceptor-mediated phosphoinositide hydrolysis.     

Phorbol Myristate Acetate Inhibition of Phospholipase C Activation by Carbachol in Slices of Rat Brain Cortex Is a Delayed and Indirect Effect

We examined the effect of phorbol esters on phospholipase C activation in rat brain cortical slices and membranes. There was little effect of concurrent addition of phorbol 12-myristate 13-acetate (PMA) with carbachol on phosphoinositide breakdown due to carbachol over a 1-h incubation of brain slices. However, if slices were preincubated for 3 h with 1 microM PMA or 200 microM sphingosine before addition of carbachol, there was a 35-50% inhibition of phosphoinositide breakdown. There was also a marked loss of protein kinase C (PKC) activity from both cytosol and membranes after a 3-h exposure to PMA. The loss in responsiveness to the muscarinic agonists in slices was not reflected in carbachol-stimulated phospholipase C activation using isolated membranes. However, the decrease in carbachol-induced phosphoinositide breakdown seen in slices after a 3-h exposure to PMA was abolished if the extracellular K+ concentration was elevated from 5.9 to 55mM. Because elevation of the K+ level induces depolarization and increases Ca2+ entry, we examined the effect of ionomycin, a Ca2+ ionophore. Ionomycin potentiated the effects of carbachol on phosphoinositide breakdown but was unable to reverse the effects of a 3-h incubation with PMA. Because apamin, an inhibitor of Ca2(+)-dependent K+ channels, mimicked the effects of exposure to PMA for 3 h, it is possible that these channels are involved in muscarinic cholinergic regulation of phosphoinositide breakdown in rat brain slices. These results support the hypothesis that prolonged PMA treatment in rat brain cortex has no direct effect on phospholipase C activation by muscarinic cholinergic stimulation.     

Brain Tryptophan Hydroxylation in the Portacaval Shunted Rat: A Hypothesis for the Regulation of Serotonin Turnover In Vivo

Regional and whole-brain tryptophan-hydroxylating activity and serotonin turnover were investigated in portacaval shunted (PCS) rats using an in vivo decarboxylase inhibition assay. To saturate tryptophan hydroxylation with amino acid substrate, rats were administered a high dose of tryptophan 1 h prior to analysis of brain tryptophan, 5-hydroxytryptophan, serotonin, and 5-hydroxyindoleacetic acid. The analysis revealed, as expected, higher brain concentrations of tryptophan and 5-hydroxyindoles and increased serotonin synthesis rate in PCS rats as compared with shamoperated controls. Saturating levels of brain tryptophan were achieved in both PCS and sham animals after exogenous tryptophan administration. The tryptophan load resulted in increased brain serotonin turnover in all regions and in whole brain compared with rats that did not receive a tryptophan load. Tryptophan-loaded PCS rats showed increased brain serotonin turnover compared with tryptophan-loaded sham rats. Regionally, this supranormal tryptophan-hydroxylating activity was most pronounced in the mesencephalon-pons followed by the cortex. It is concluded that, at least in the PCS rat, brain tryptophan hydroxylation is an inducible process. Since it is known that brain tissue from PCS rats undergoes a redox shift toward a reduced state and that the essential cofactor tetrahydrobiopterin is active in tryptophan hydroxylation only when present in its reduced form, it is hypothesized that this is the reason for the supranormal tryptophan-hydroxylating activity displayed by the PCS rats. The hypothesis further suggests that alterations in tetrahydrobiopterin availability may serve as a mechanism by which brain tryptophan hydroxylation, and therefore serotonin turnover, can be regulated with high sensitivity in vivo.