Importance of central catecholamines in the mediation of lordosis behaviour in ovariectomized rats treated with estrogen and inhibitors of monoamine synthesis

Summary The effect of-methyl-p-tyrosine methylester hydrochloride (-MT) (100 mg/kg i.p.) orp-chlorophenylalanine methylester hydrochloride (PCPA) (150 mg/kg i.p.) on lordosis behaviour in estrogen pretreated spayed rats was investigated. The effect of-MT was studied 1, 2, 4, 8 and 26 hours after the drug injection and the observations after PCPA were done 2, 4, 8, 26 and 50 hours after the injection. Facilitation of lordosis was seen 2–8 hours after both treatments. Biochemical experiments were done in parallel and the injection of either-MT or PCPA resulted in a transient decrease in brain catecholamines which was correlated in time with the facilitation of the lordosis response. Furthermore PCPA resulted in a gradual decline in brain 5-HT. However, when 5-HT depletion was most pronounced and the brain catecholamines had returned to control levels no increase in lordosis behaviour occurred. Our findings suggest a role for central catecholamines in the mediation of lordosis behaviour in ovariectomized estrogen-drug-treated rats.     

Dose-dependent pharmacokinetics of α-methyl-p-tyrosine (α-MT) and comparison of catecholamine turnover rates after two doses of α-MT

Summary Groups of rats were injected i.p. with 0.407 or 1.02 mmoles/kg of D, L--methyl-p-tyrosine methylester HCl (-MT). The time-courses for-MT in plasma and brain were followed together with the endogenous brain dopamine (DA) and noradrenaline (NA) contents.The elimination of-MT from plasma and brain was markedly delayed after the high-MT dose compared with the low dose. At 40 hours after the injection of 1.02 mmoles/kg of-MT both plasma and brain levels were high, whereas no-MT could be detected in plasma or brain at 16 hours after the lower dose.The brain catecholamines were decreased to very low values after the higher-MT dose (DA 14% and NA 10% of controls at 8 and 24 hours respectively). There was no complete recuperation at 40 hours of any of the amines. After the lower-MT dose, the DA concentration was back to control levels at 16 hours and NA at 12 hours. Between 16–40 hours after the high-MT dose a majority of the rats showed prominent signs of sedation, weight loss and dehydration. No such signs were observed in rats receiving 0.407 mmoles/kg. During the first hour after the-MT injection the declines of DA and NA respectively were almost identical for both-MT doses. When the whole time-course (0–8 hours) after the high dose was considered, biphasic declines were obtained for both DA and NA, suggesting at least two different catecholamine pools. However, due to toxic effects after the high-MT dose, turnover data have to be interpreted with caution.     

Lack of effect of a behaviorally active dose ofα-melanotropin on biochemical indices of dopaminergic neuronal activity in the rat

Summary An attempt was made to correlate behavioral changes in the male rat following an intracerebroventricular (ICV) injection of-melanotropin (-MSH, 10g) with biochemical estimates of the activities of nigrostriatal, mesolimbic, tuberoinfundibular and tuberohypophyseal dopaminergic neurons. ICV injection of-MSH elicited body snaking, stretching, yawning and penile erections. The concentrations of dopamine (DA) and dihydroxyphenylacetic acid and the rate of DA synthesis (accumulation of DOPA after the inhibition of DOPA decarboxylase) in the striatum, nucleus accumbens, olfactory tubercle, septum, median eminence and posterior pituitary were unaltered at any time (30–180 min) after ICV injection of-MSH. Intraperitoneal injection of-MSH (100g/kg) also failed to change the rate of DOPA accumulation in these brain regions. These results imply that the behaviors observed after-MSH are not associated with changes in the activity of DA neurons.Visiting professor from Fukuoka University, Fukuoka, Japan.     

Activation of α2-adrenoreceptors enhances haloperidol-induced suppression of operant behavior

Summary Inhibition of catecholamine synthesis by-methyl paratyrosine (-MT) was previously shown to potentiate the behavioral suppression caused by dopamine-receptor antagonists. This effect of-MT is in all probability due to inhibition of the compensatory increase in dopamine turnover induced by the dopamine receptor antagonists. In the present study we investigated the effect of the ₂-adrenoreceptor agonist clonidine on the haloperidol-induced suppression of food-reinforced lever-pressing behavior (fixed ratio 401) in rats. Small behaviorally inactive doses of clonidine were found, in analogy with-MT, to enhance the haloperidol-induced suppression of the lever-pressing behavior. The haloperidol-induced increase in dopamine synthesis (measured as the accumulation of DOPA after inhibition of aromatic amino acid decarboxylare) was antagonized by clonidine in the striatum as well as in the dopamine rich limbic regions. Prazosin, a selective ₁-adrenoreceptor antagonist had no effect on the clonidine induced behavioral changes. Idazoxane, a selective ₂-adrenoreceptor antagonist, counteracted both the behavioral and biochemical effects of clonidine, indicating that these effects of clonidine are mediated via its action on ₂-adrenoreceptors. The present findings provide support for the notion that ₂-adrenoreceptors may participate in the regulation of nigro-striatal as well as meso-limbic dopaminergic activity. It is suggested that ₂-adrenoreceptor agents, especially in combination with classical antipsychotics, might be of therapeutic value in the treatment of disorders associated with abnormal dopaminergic activity.     

Time course changes of estrogen receptor α expression in the adult rat hippocampus after kainic acid-induced status epilepticus

Although estrogens possess neuroprotective and epileptogenic properties, the expression pattern of the estrogen receptor (ER) following status epilepticus (SE) remains unclear. We therefore examined the expression pattern of ER in the adult rat hippocampus after SE. SE was induced in rats by kainic acid (KA; 12 mg/kg, i.p.). ER expression was assessed by immunostaining and Western blotting at various times (24 h, and 7, 14, and 21 days) after SE onset. Immunohistochemistry disclosed ER expression in the CA1 and CA3 pyramidal cells of control rats, whereas, after SE, ER-immunoreactive neurons decreased in number due to neuronal death in the CA1 from days 7 to 21. On the other hand, ER-immunoreactive cells with astrocytic morphology were observed in the CA1 beginning on day 7 after SE. This immunoreactivity increased in proportion to the hypertrophy of astrocytes up to day 21. Western blotting revealed a significant decrease in ER expression on day 7 after SE in comparison with control level. However, ER expression on days 14 and 21 were similar when comparing KA-treated and control rats. These results indicate that reactive astrocytes are important sites of estrogen action in the hippocampal CA1 after SE.     

Histopathological effects of intracerebral injections of human recombinant tumor necrosis factor-α in the rat

Summary Human recombinant tumor necrosis factor- (rTNF-) was administered to normal Fischer 344 rats by stereotaxic intracerebral (IC) injection. Animals received a single injection of either 6×10⁴ UrTNF- or excipient in their right parietal lobe. Others received three consecutive daily injections of either 6×10⁴ U rTNF- or excipient to examine effects of higher accumulative doses. Histological examination of the brain revealed that both single and multiple IC injections of rTNF- triggered an immigration of circulating leukocytes into the site of TNF- injection. After one injection, this cell population was composed mainly of macrophages and neutrophils. Maximal leukocytic influx occurred by 48 h and was composed mostly of neutrophils which were limited to the injection site and perivascular space. Quantitation of the inflammatory reaction by measurement of tissue myeloperoxidase levels supported these histological observations. One day after multiple rTNF- injections, leukocytic adhesion to endothelium, vascular cuffing and leukocytic infiltration into the neuropil was observed at levels comparable to those seen 3 days following a single rTNF- injection. We conclude that while one or more IC injection(s) of 6×10⁴ U rTNF- was well tolerated in normal rats, at this dose the cytokine triggers a pronounced leukocytic infiltration at the site of injection. These results support a role for TNF- as a mediator in inflammatory responses within the central nervous system.Supported in part by a grant from the A. D. Williams Research Fund and by gifts from the Kellogg Foundation, the Lind Lawrence Fund, and the family and friends of Christine Armstrong, Jack Harvey, Christopher Wemple, and Pearl Ylonen.     

Antipsychotic treatment withα-methyltyrosine in combination with thioridazine: Prolactin response and interaction with dopaminergic precursor pools

Summary The antipsychotic effect of-methyltyrosine (-MT) in combination with thioridazine was investigated by means of rating scales for social behaviour and mental symptoms The clinical effect was also evaluated in relation to the serum concentrations of-MT and thioridazine and to the increase in prolactin secretion in response to the interaction with hypothalamic dopaminergic mechanisms. The interactions between the serum levels of-MT and those of the transmitter precursors phenylalanine and tyrosine were analysed. The results confirmed the ability of-MT (2g/day) to potentiate the antipsychotic effect of thioridazine, whereby the dose of neuroleptic drug required to control psychotic symptoms may be markedly reduced. None of the four patients who completed the trial showed side effects that could be ascribed to-MT. The antipsychotic effect of thioridazine, alone or in combination with-MT, correlated well with the prolactin response in the individual patient. No important interference with serum phenylalanine or tyrosine levels was noted during treatment with-MT.     

Effect of clonidine on ultrasonic vocalization in preweaning rats

Summary The present study was undertaken to investigate the involvement of the noradrenergic neurotransmission system in the ultra sonic callings emitted by rat pups separated from their mother and exposed to cold stimulation. The investigation was primarily performed by help of agents selectively affecting the -adrenoceptors: the ₂-agonist clonidine, the ₁-antagonist prazosin and the ₂-antagonist idazoxan.Clonidine dose-dependently stimulated the amount of ultra sonic vocalization, an effect not solely dependent upon the effect of clonidine on body temperature. In a developmental study it was found that clonidine uniformly stimulated crying at all ages from 4 days of age up to 18 days of age, that is during the whole preweaning period. Clonidine stimulated ultrasonic crying in rat pups, devoid of presynaptic catecholamine (CA) neurons by combined pretreatment with the monoamine depletor, reserpine, and the inhibitor of CAsynthesis, -methyl-tyrosine. This finding suggested that the stimulating effect of clonidine on ultrasonic vocalization was mediated by postsynaptic adrenoceptors.In pups, 12 days of age, idazoxan blocked the effect of cold stimulation on ultra sonic crying, suggesting that ₂-adrenoceptors, presumably postsynaptic ones, mediated this kind of stimulation. Idazoxan also antagonized the effect of clonidine, but only at a dose effective also in control pups. Prazosin had no effect on cold-stimulated crying, but antagonized the effect of clonidine, suggesting that the effect of clonidine was also mediated by ₁-receptors. At 18 days of age, prazosin no longer antagonized the effect of clonidine, whereas the antagonizing action of idazoxan was reinforced.The age-dependent variation in responsiveness to the adrenergic drugs suggest maturational changes in the function of the CA-system occurring between 12–16 days of age.     

On areas of transition between entorhinal allocortex and temporal isocortex in the human brain. Normal morphology and lamina-specific pathology in Alzheimer's disease

Summary The allocortical entorhinal region does not gradually transform into the temporal isocortex. Instead, there is an extended stretch of transentorhinal cortex with interdigitation of allocortical and isocortical laminae. The main feature of this transition zone is that the superficial layer of large multipolar nerve cells (Pre-) of the entorhinal region gradually sweeps downward and follows an oblique course through the outer layers. During this course the starshaped nerve cells of Pre- are transformed into pyramidal cells.The layer Pre- projection cells are particularly prone to the development of neurofibrillary changes of the Alzheimer type. In cases of presenile and senile dementia almost all of the layer Pre- projection neurons are changed pathologically. The isocortical pyramidal cells of layers II to IV are far less inclined to develop neurofibrillary changes. In the transentorhinal cortex, the tangle-bearing neurons follow an oblique course through the superficial laminae and are finally located between the isocortical layers III and IV, findings that confirm the assumption that these neurons are constituents of the allocortical layer Pre-.Layer-specific pathology of the profound stratum as well confirms the transentorhinal region as being formed by interdigitating allocortical and isocortical layers.Supported by grants from the Deutsche Forschungsgemeinschaft     

αB-Crystallin is present in reactive glia in Creutzfeldt-Jakob disease

Summary -Crystallin is a major eye lens protein, composed of two types of subunits, A and B. The A subunit is restricted to the lens, but B-crystallin has recently also been detected in non-lenticular tissues, including the nervous system. With the use of a polyclonal antiserum directed against a synthetic C-terminal peptide of human B-crystallin, the presence of B-crystallin could be demonstrated immunohistochemically in astrocytes in the brains of patients with Creutzfeldt-Jakob disease (CJD). Most intensive localization was observed in the spongiotic tissue representing abundant progressively changed astrocytes in CJD. In agematched control brains weak positive reaction was located in individual oligodendroglia cells and subpial astrocytes. Prominent increase of B-crystallin in pathological glia in CJD may represent a response to stress.     

The effect of β-alanine on motor behaviour,body temperature and cerebral monoamine metabolism in rat

Summary Intracerebroventricular (ICV) injection of-alanine produced a decrease in rectal temperature, inhibition of exploratory behaviour and motility, and changes in the metabolism of cerebral monoamines. Dopa and 5-HTP accumulation after inhibition of L-aromatic amino acid decarboxylase, NSD 1015 (3-hydroxybencylhydrazine HCl, 100mg/kg i.p.) was found to be significantly increased in all the dissected cerebral regions of animals treated with-alanine, as compared to the controls. Levels of tyrosine and tryptophan did not show any significant change. Endogenous levels of dopamine (DA), noradrenaline (NA), serotonin (5-HT) and 5-hydroxyindolacetic acid (5-HIAA), did not change. After inhibition of the catecholamines synthesis with-methyltyrosine (-MT), dopamine depletion was retarded and noradrenaline accelerated, but without reaching statistical significance. After intraperitoneal (i.p.) injection of-alanine, significant changes in motor behaviour were found. Body temperature and metabolism of brain catecholamine were unchanged. This lack of effect could be explained by poor penetration through the blood-brain barrier.     

The hyperkinetic syndrome following long-term haloperidol treatment: Involvement of dopamine and noradrenaline

Summary Mice withdrawn for 7 days from a 35-day treatment period with haloperidol (3 mg/kg/day) displayed significantly greater spontaneous locomotor activity (hyperkinesia) than animals withdrawn from the vehicle. The hyperkinesia was antagonized by phenoxybenzamine (an-adrenergic receptor antagonist) and by FLA-63 (a dopamine--hydroxylase inhibitor) but not by haloperidol (a dopamine receptor antagonist).-Methyl tyrosine (a tyrosine hydroxylase inhibitor) was effective in antagonizing the hyperkinesia and this blockade by-methyl tyrosine could be completely reversed by the administration of a low dose of the catecholamine precursor, DOPA. The data suggest that noradrenergic systems are of importance for the manifestation of the hyperkinetic syndrome seen in mice withdrawn from long-term haloperidol treatment.     

Effect ofd,l-α-aminoadipate on the mediobasal hypothalamus and endocrine function in the rat

Summary Using the glutamate analog,d,l--aminoadipic acid (d,l-AA), experiments were conducted to examine the nature, extent, and specificity of its toxicity in the mediobasal hypothalamus and to determine its effect on endocrine homeostasis. Neonatal rats received daily injections ofd,l-AA (4 g/kg BW) on postnatal days 5–10 and were killed at various post-treatment intervals. Sex-matched littermates were given equimolar amounts of NaCl and served as controls. Treated rats killed 18 days post injection weighed slightly less than controls and had reduced testicular, ovarian, and uterine weights, but the differences were not statistically significant. Ind,l-AA treated rats serum and pituitary levels of TSH and PRL were comparable to control values. Pituitary content of LH ('s and 's) and FSH ('s), however, was lower (P<0.05) ind,l-AA treated rats than in controls, but serum levels were not significantly different. Distinct cytopathologic changes were evident in the arcuate nucleus and median eminence ofd,l-AA-treated rats killed at 2 and 6 h post injection only. By 12 h evidence of acute damage had largely disappeared. Both glial and ependymal cells underwent edematous swelling and necrosis, but neurons were largely unaffected. Evidence of reactive changes, such as gliosis, infiltration of microglia, and removal of debris, however, were not very conspicious. A random sample of mediobasal hypothalami of rats killed 18 days post injection failed to show any detectable lesion or residual effects of earlier pathology. Age at the time of exposure to the gliotoxin was found to be an important variable affecting both extent and duration of injury. The most deleterious effects were observed when the gliotoxin was administered in the form of a single injection on postnatal day 5 only. The results suggest that normal neuronal activity and endocrine homeostasis, specifically gonadotropin, may be irreversibly altered as a consequence of transient disruption of the glial compartment.Supported by grants from the Medical Research Council of Canada, the St. Boniface General Hospital, and Mrs. James A. Richardson Research Foundations     

Increasedα 2;-adrenoceptor density in the frontal cortex of depressed suicide victims

Summary The density of brain ₂-adrenoceptors, quantitated by means of the binding of the agonist [³H]clonidine, was studied in post-mortem cortical membranes of matched control subjects and depressed suicide victims. In the depressed suicide group, the specific high affinity binding of [³H]clonidine was found to be significantly increased (B_max, 72% greater; p<0.01) without significant changes in the K_D value for the radioligand. These preliminary results indicate that ₂-adrenoceptor density in the high affinity state _2H) is increased in the brain of depressed patients and add strong support to the hypothesis that endogenous depression is related to supersensitive ₂-adrenoceptors.     

αB-crystallin in oxidative muscle fibers and its accumulation in ragged-red fibers: a comparative immunohistochemical and histochemical study in human skeletal muscle

Summary The B-crystallin gene is abundantly experssed in the vertebrate lens and at lower levels in various non-lenticular tissues. Among the non-lenticular tissues, B-crystallin is present at high levels in the heart and skeletal muscle. Using a specific antibody against B-crystallin, the cellular localization of B-crystallin was studied in biopsies of human skeletal muscles. Expression of B-crystallin was observed in normal oxidative muscle fibers that show positive reactions for NADH-tetrazolium reductase and cytochrome c oxidase. In muscle diseases increased immunoreactivity for B-crystallin was found in ragged-red fibers, which stained darkly with histochemistry for succinate dehydrogenase. Since B-crystallin is related to small heat-shock proteins and can be induced by various stress conditions, the increased B-crystallin immunoreactivity of ragged-red fibers could result from profound oxidative stress produced by the abnormal mitochondrial metabolism.Supported by NIH grants NS 17125 and EYO9331 (J.E.G.)     

Effects of chlorimipramine and protriptyline on the hyperactivity induced by 5-hydroxytryptophan after peripheral decarboxylase inhibition in mice

Summary DL-5-HTP in doses from 100 mg/kg is reported to increase the spontaneous motor activity in mice pretreated with L--hydrazino--methyl--(3, 4-dihydroxyphenyl)-propionic acid (MK-486), an inhibitor of peripheral aromatic aminoacid decarboxylase. Exogenously administered 5-HTP gives rise to accumulation of 5-HT in both central 5-HT-and catecholamine-neurons. The stimulant effect of DL-5-HTP may be mediated via increased activation of 5-HT receptors or, via displacement, increased activation of catecholamine receptors. In the present experiment pre-treatment with chlorimipramine, 25 mg/kg concomitant with MK-486 markedly potentiated the stimulant effect of DL-5-HTP on motor activity,i.e. the dose response curve was shifted to the left. DL-5-HTP, 75 mg/kg, induced in these animals a pronounced increase of the motor activity but had no significant effect on the brain concentrations of DA or NA. Pre-treatment with protriptyline-HCl, 25 mg/kg, concomitant with MK-486 did not potentiate the stimulant effect of DL-5-HTP. The results indicate that the DL-5-HTP induced motor activity is largely dependent on an increased activation of central 5-HT receptors.     

Colocalization of growth hormone (GH) and glycoprotein subunit α in GH-producing pituitary adenomas in acromegalic patients

Thirty-one consecutive cases of pituitary adenoma in acromegalic patients were studied by immunohistochemistry. All adenomas contained cells immunoreactive with the anti--subunit of gonadotropic hormones (; 0.6–53% of tumor cells) as well as with anti-growth hormone (GH; 4–74% of tumor cells). In serial section study, most cells immunoreactive with anti- were identical to cells immunoreactive with anti-GH. There was a positive correlation between the percentages of cells immunoreactive for in GH cells [(%)/GH(%)] and those for prolactin (PRL) in immunoreactive tumor cells {PRL(%)/[PRL(%)+GH(%)]} in mixed GH cell-PRL cell adenomas, suggesting that the -subunit may play a role in emergence of PRL cells.     

Expression of αB-crystallin in Alzheimer's disease

B-crystallin is a member of the small heatshock protein family. Under pathological conditions, the expression of B-crystallin increases in proliferating astrocytes, which suggests that this protein, in addition to glial fibrillary acidic protein (GFAP), can be a marker for gliosis in neurodegenerative diseases. Immunoblotting and immunohistochemical methods were used for the detection of B-crystallin in the brains of Alzheimer's disease (AD) patients and nondemented controls. An increase in B-cyrstallin expression was found in the brains of AD patients. Immunoreaction was present in reactive astrocytes, microglia, and oligodendrocytes, indicating that all types of glia respond to the stress associated with AD pathology. Colocalization of GFAP and B-crystallin was found in fibrous astrocytes. However, the intensity and range of B-crystallin expression appeared to be limited as compared with the large increase in the number of GFAP-positive astrocytes. This indicates that expression of B-crystallin is not a marker for gliosis in AD. Immunoreactivity to B-crystallin in both astrocytes and microglia was found mainly restricted to areas with senile plaques and neurofibrillary tangles, suggesting the association of B-crystallin with amyloid deposition in AD.Supported by a grant (No. EY08202) from the National Institutes of Health, Bethesda, USASupported by a fellowship of the Royal Netherlands Academy of Arts and Sciences     

Attenuation of contractile responses to sympathetic co-transmitters in veins from subjects with essential hypertension

Neuropeptide Y (NPY), noradrenaline (NA) and ATP are cotransmitters of the sympathetic nervous system and exert vasocontractile effects. The aim of this study was to determine the role of these sympathetic co-transmitters in human hypertension. Subcutaneous vessels from 12 patients with essential hypertention and 12 matched controls were studiedin vitro. Vascular contractile responses to NPY, NA, ,-methylene ATP (,-mATP) and potassium were studied in isolated arteries and veins (diameter 0.1–1.1 mm) with intact endothelium. The dilatory effect of acetylcholine was used to test the endothelial function. There was no difference in potency (pD₂) or contractile response to NPY, NA or ,-mATP between hypertensive and control arteries. In veins, however, the contractile response to NPY was signficantly reduced in hypertensives and the responses to NA were unchanged. Furthermore, the sensitivity (pD₂) to ,-mATP was significantly reduced in veins from hypertensives. There was no difference in the dilatory response to acetylcholine between the hypertensives and the controls, neither in the arteries nor in the veins, indicating that the observed changes in vascular reactivity to NPY, NA and ,-mATP were not endothelium-dependent. In conclusion, the postjunctional contractile effect of NPY and sensitivity (pD₂) to ,-mATP, co-transmitters of the peripheral sympathetic nervous system, are attenuated in veins in essential hypertension.     

Tumor necrosis factor alpha serum levels and inflammatory response in acute ischemic stroke patients

Abstract. Experimental evidence indicates that tumor necrosis factor alpha (TNF-) is involved in brain damage following ischemic injury. The present study was designed to monitor serum TNF- levels in acute stroke patients and to correlate TNF- levels with lesion size, neurological impairment and vascular risk factors. In 41 patients with ischemic stroke, serum TNF- levels were serially measured by a solid enzyme amplified sensitivity immunoassay (EASIA) in the first 10 days after stroke onset. Serum fibrinogen and Creactive protein (CRP), white blood cell (WBC) and neutrophil counts were determined on the same days to monitor acute phase response changes. Lesion size was calculated on computed tomograms by a computer-assisted procedure. Neurological impairment was evaluated on the Canadian Neurological Scale. Forty age-matched subjects were used as controls. Compared to baseline, TNF- levels significantly increased during the study (p=0.0001), peaking on day 7. Peak TNF- levels did not correlate with neurological impairment or lesion size. Multivariate analysis showed that sex, age, vascular risk factors and infectious complications did not influence TNF- levels. Fibrinogen, CRP, WBC and neutrophil concentrations increased, indicating an acute phase response occurred after stroke. In conclusion, serum TNF- levels showed an early and prolonged increase after stroke onset, unrelated to lesion size, neurological impairment, age, sex, vascular risk factors or infectious complications. Serum increase of TNF- may be explained as part of the acute phase response occurring in stroke patients.