Content of endogenous opiates and adrenocorticotropic hormone in the brain structures of rats of different ages

The contents of beta-endorphin (BE), methionine-enkephalin (MEK), and adrenocorticotropic hormone (ACTH) in the hypophysis and hypothalamus of intact 4- to 6-week-old and 16-week-old Wistar rats was studied. The maximum BE concentration was found in the hypophysis, whereas the maximum MEK and ACTH concentrations were found in the hypothalamus. Aging was followed by a decrease in the concentrations of all above substances, except BE, whose concentration in the hypophysis of the older rat group was markedly higher than in the hypophysis of 4- to 6-week-old animals.     

Effect of the local administration of 5,7-DHT and 6-OHDA into the neocortex on the learning and exploratory behavior of rats in an open field

On Wistar rats characteristics were studied of investigating behaviour in the open field, of learning of conditioned food-reinforced reaction and also of BA and their metabolites content in various brain structures under local intracerebral injections of specific neurotoxins; 6-hydroxydopamine (6-OHDA) and 5,7-dihydroxytryptamine (5,7-DHT), abolishing correspondingly catecholaminergic and serotoninergic terminals. Bilateral injection of 6-OHDA in the neocortex led to a weakening of rats investigating activity in the open field and to an increase of the time of fulfillment of the forming of conditioned food-reinforced reaction. Administration of 5,7-DHT was accompanied by an increase of the investigating behaviour in the open field and a reduction of the duration of the forming of conditioned reaction. Administration of 6-OHDA to the neocortex caused a lowering of catecholamines level in the frontal area of the neocortex and the hippocampus. Analogous administration of 5,7-DHT elicited simultaneously with a deep level lowering of 5-HT and its metabolite in these structures, a change of catecholamines content which testifies to a lesser specificity of the neurotoxin 5,7-DHT in comparison with 6-OHDA. Structures lesion of serotoninergic and catecholaminergic systems of the frontal cortex and the hippocampus brought about by a local administration of 6-OHDA and 5,7-DHT in the neocortex was accompanied by differently directed changes in animals behaviour.     

Changes in the hypothalamus, reticular formation and pituitary of rats during development of subcutaneous tumors]

Dynamics of structural changes was studied in the hypothalamus, reticular formation and the hypophysis of rats, depending on the phases of development in them of 9m10-dimethyl-1,2-benzanthracene-induced tumours. The phase of subcutaneous tumours appearance proved to be accompanied by structural and functional changes in the hypothalamus, reticular formation and the hypophysis.     

Effects of gonadectomy and sex hormones on the levels of noradrenaline and dopamine in rat hypothalamus

The levels of noradrenaline and dopamine were determined in the homogenates of the hypothalamus of rats of either sex. The determinations were done in intact rats, after sham gonadectomy, 6 and 9 weeks after gonadectomy, and in gonadectomized rats receiving 6 weeks after gonadectomy one dose of oestradiol cypionate (females) or testosterone cypionate (males). Catecholamines were determined fluorimetrically. The changes of the determined catecholamines differed in the hypothalamic homogenates in males and females after gonadectomy. Following orchidectomy the noradrenaline level rose, while after ovariectomy the level of this catecholamine decreased. Contrary to this, in ovariectomized rats the dopamine level was significantly raised after the operation. This change was reversible as observed after administration of oestradiol cypionate. Orchidectomy and testosterone cypionate injection had no effect on the dopamine level in the hypothalamus. The role of these catecholamines in the processes connected with the regulation of the hypothalamus-hypophysis-gonad axis is discussed.     

Differential Effects of Chemical Sympathectomy on Expression and Activity of Tyrosine Hydroxylase and Levels of Catecholamines and DOPA in Peripheral Tissues of Rats

Tyrosine hydroxylase (TH) mRNA and activity and concentrations of 3,4-dihydroxyphenylalanine (DOPA) and catecholamines were examined as markers of sympathetic innervation and catecholamine synthesis in peripheral tissues of sympathectomized and intact rats. Chemical sympathectomy with 6-hydroxydopamine (6-OHDA) markedly decreased norepinephrine and to a generally lesser extent TH activities and dopamine in most peripheral tissues (stomach, lung, testis, duodenum, pancreas, salivary gland, spleen, heart, kidney, thymus). Superior cervical ganglia, adrenals and descending aorta were unaffected and vas deferens showed a large 92% decrease in norepinephrine, but only a small 38% decrease in TH activity after 6-OHDA. Presence of chromaffin cells or neuronal cell bodies in these latter tissues, indicated by consistent expression of TH mRNA, explained the relative resistance of these tissues to 6-OHDA. Stomach also showed consistent expression of TH mRNA before, but not after 6-OHDA, suggesting that catecholamine synthesizing cells in gastric tissue are sensitive to the toxic effects of 6-OHDA. Tissue concentrations of DOPA were mainly unaffected by 6-OHDA, indicating that much of the DOPA in peripheral tissues is synthesized independently of local TH or sympathetic innervation. The differential effects of chemical sympathectomy on tissue catecholamines, DOPA, TH mRNA and TH activity demonstrate that these variables are not simple markers of sympathetic innervation or catecholamine synthesis. Other factors, including presence of neuronal cell bodies, parenchymal chromaffin cells, non-neuronal sites of catecholamine synthesis and alternative sources of tissue DOPA, must also be considered when tissue catecholamines, DOPA and TH are examined as markers of sympathetic innervation and local catecholamine synthesis.     

Effects of 6-hydroxydopamine on brain and blood catecholamines, ammonia, and amino acids in rats

The effect of 6-hydroxydopamine (6-OHDA) upon brain and blood catecholamines, ammonia, and amino acids has been studied in rats subjected to increasing doses of the drug. Time dependent effects after injection have also been studied. Systemically injected 6-OHDA significantly, acutely reduced brain adrenaline (A), noradrenaline (NA), total catecholamines (TC), gamma-aminobutyric acid (GABA), and glutamic acid (Glu); concomitantly brain ammonia (NH3) increased. In blood, NA and TC were reduced and A and NH3 increased. The changes in brain monoamines are surprising since it has been reported that 6-OHDA does not cross the blood-brain barrier. We have proposed that these changes result from a general stress response or a reflex peripheral sympathetic response to falling blood pressure which in some manner communicates to the central nervous system. As the dose of 6-OHDA increased, brain NH3 increased and Glu decreased. A similar effect was seen from a single dose as the time after injection for sampling brain and blood constituents increased. Blood ammonia increases without change in Glu, glutamine, or asparagine. The source of NH3 may be from deamination of adenine nucleotide or catechols released from nerve terminals under the abnormal stimulus of 6-OHDA.     

Effects of hypothalamic microinjections of 6-hydroxydopamine (6-OHDA) on estral cycle and morphology of the genital tract in the female rat (author's transl)]

To determine whether central catecholaminergic pathways are involved in the neural contral of gonadotrophin secretion, they were interrupted at the hypothalamic level by microinjections of 6-hydroxydopamine (6-OHDA). The effects on ovulation, estral cycle and ovarian and uterine histology were studied. Microinjections of 50 mug of 6-OHDA hydrobromyde were made bilaterally into the anterolateral hypothalamus in a group of rats. Another group was injected with 25 mug of 6-OHDA, while a control group recieved an equivalent volume (5 mul) of saline with ascorbic acid. Animals injected with 50 mug of 6-OHDA showed blockade of ovulation, vaginal cytology characteristics of persistent estrous, polyfollicular ovaries and enlarged uteri with hypertrophic endometrial glands. In the group injected with 25 mug, similiar effects were demonstrated, but the number of affected animals was smaller than that in the 50 mug group. Control animals dit not show modifications, either in estral cycle or in ovarian and uterine histology. These results suggest that 6-OHDA injected into the anterolateral hypothalmus interferes with catecholaminergic pathways that participate in the neural control of ovulation.     

Effects of endogenous opioids on the development of reproductive function in rats]

The influence of opioid peptide on the process of formation of reproductive function in rats was studied. Administration of beta-endorphin to neonatal female rats did not affect the concentrations of oestrogen and androgen receptors in the hypothalamus and pituitary, whereas the content of testosterone receptors was significantly higher in both hypothalamus and pituitary. Chronic administration of beta-endorphin to both female and male rats does not affect the concentration of sex hormones. The results obtained indicate that chronic administration of beta-endorphin to neonatal female rats lead to formation of instable contacts in the mechanism of regulation of hypophysis gonadotropic function.     

Lesions of the ventral noradrenergic bundle prevent the rise in blood pressure induced by social deprivation stress in the rat

1. Hypertension can be induced by some types of stress in the rat. The aim of the present work was to study the putative implication of brain norepinephrine (NE) in blood pressure increase due to social deprivation stress. 2. The effects of 6-hydroxydopamine (6-OHDA) lesions of the ventral noradrenergic bundle (VNEB) on the hypertensive response induced by brief social deprivation stress in young Wistar rats were examined. NE, dopamine (DA), and epinephrine (EPI) levels were measured by HPLC coupled with electrochemical detection in two brain areas (hypothalamus and medulla oblongata) relevant for blood pressure regulation. 3. VNEB lesions prevented the hypertensive response produced by isolation. Twelve or 20 days after 6-OHDA administration, NE and EPI but not DA levels decreased in the hypothalamus of the lesioned rats. In contrast, no catecholamine changes were detected in medulla oblongata. 4. These data suggest that the VNEB plays a role in the triggering of the hypertensive response induced by social deprivation stress in young Wistar rats.     

A possible role for spinal noradrenaline in the mechanisms of 6-hydroxydopamine against pentylenetetrazol induced convulsions in rats

The threshold of the generalized clonic convulsions induced by intravenous infusion of pentylenetetrazol (PTZ) was significantly increased by the intraperitoneal administration of noradrenaline (NA) neurotoxin, 6-hydroxydopamine, which produced no changes in the levels of catecholamines in discrete areas of rat brain, but the effect was accompanied by spinal depletion of NA. Moreover, the anticonvulsant effects of phenobarbitone (PB) and diphenylhydantoin (DPH) against PTZ convulsions were also significantly increased in the animals pretreated with 6-OHDA. These results suggest that the observed elevation of PTZ convulsive threshold and the potentiation of anticonvulsant activity of PB and DPH in 6-OHDA treated rats were possibly mediated through spinal cord depletion of NA.     

Secretory and structural effects of 6-hydroxy-dopamine on normal parotid glands of rats, and at different times after surgical sympathectomy.

The effects of i.v. 6-hydroxydopamine (6-OHDA), 100 mg/kg, have been studied on parotid glands of rats at 12, 24, 48, 72 hr and 3 weeks after avulsion of the right superior cervical sympathetic ganglion. The salivary flow from normal left control glands and from right glands 12 hr after ganglionectomy were similar, but at longer times after ganglionectomy the secretory response from the test glands was greatly reduced. Morphological assessment showed that 6-OHDA induced a massive depletion of secretory granules from all control glands and also at 12 hr after ganglionectomy but at 48 and 72 hr there was considerably less depletion of granules on the ganglionectomized side. It is thought that at the longer times after ganglionectomy the secretion from the test glands is caused by circulating catecholamines released by the action of 6-OHDS on adrenergic nerves elsewhere, plus a possible small direct secretogogue effect oomy are thought to be attributable to the release of catecholamines from adrenergic nerves within the gland.     

Evidence of an epididymis factor inhibiting the hypothalamic synthesis of FSH-RH in the rat]

We have measured, by radioimmunoassay, FSH and LH in the blood plasma and in the hypophysis of castrated male rats, injected with epididymal inhibin; we have also evaluated the FSH and LH releasing activities of their hypothalamus by measuring plasma FSH and LH levels of spayed female rats, treated by hypothalamic extracts of the previous rats. The FSH and LH pituitary levels do not change compared with controls, and it is impossible to know if inhibin acts directly on hypophysis; it is likely that, directly or indirectly, inhibin restrains at the same time the synthesis and the release of FSH. On the contrary, the hypothalamic extracts lose their FSH-RH, but not their LH-RH, activities; then, inhibin operates on hypothalamus by suppressing of the synthesis of FSH-RH.     

Dopamine sensitive neurons of the arcuate region of the hypothalamus and their role in regulating the gonadotropic function of the pituitary]

The effect of the microiontophoretic application of dopamine (DA) into the arcuate region of the hypothalamus on the sensitivity of single neurons and on the plasma and hypophysis LH levels was examined at various stages of the estrous cycle. During the estrous cycle in rats there was no significant differences in the relative number of neurons showing activation and inhibition or non-responsive to DA. However, in the first half of proestrus (P) a significant increase in the number of neurons with the excitative reaction to the iontophoretic application of DA was observed. At all the stages of the cycle, except the second half of P, the excitative reaction of neurons correlated with increased LH level in the plasma. In the hypophysis only in diestrus-2 there was a significant increase of the LH level in response to the iontophoretic application of DA in the arcuate neurons of the hypothalamus.     

Functional maturation of the human corticotropin releasing factor-adrenocorticotropic hormone system during intrauterine life. An in vitro study

Radioimmunoassay was used to determine ACTH secretion by cultured hypophyses of human fetuses from the 6th to the 30th week of intrauterine life and their responsiveness to hypothalamic extracts obtained from adult animals. CRF-like activity in the human hypothalamus was measured within the 6th to the 32nd week of prenatal development from changes in ACTH release by cultured cells of the adult rat hypophysis. It was established that starting from weeks 6-7 of embryogenesis, the human fetal hypophysis is capable of synthesizing and secreting immuno-reactive ACTH in vitro. The human fetus hypothalamus of the first trimester of gestation contained no CRF-like substance. The fetus hypothalamus of the second and third trimesters of pregnancy manifested a considerable amount of CRF-like substance. It is suggested that CRF appears at the end of the first trimester of pregnancy.     

The effects of reserpine and haloperidol on tyrosine hydroxylase activity in the brains of aged rats

Many neurotransmitter systems appear to be altered with aging. The effects of aging on the regulation of tyrosine hydroxylase, the rate-limiting enzyme in the synthesis of catecholamines in the brain has been examined. The endogenous basal activity of tyrosine hydroxylase was lower in the hypothalamus of 24 month old Fisher 344 rats than in the hypothalamus of 3 month old or 6 month old animals. There was no difference in the basal activity of tyrosine hydroxylase in the locus ceruleus, frontal cortex, hippocampus, substantia nigra, or the striatum of rats of ages 3 months, 6 months and 24 months. Tyrosine hydroxylase activity was increased in the striatum of 3 month old (60%) and 6 month old (28%) rats after treatment with haloperidol or reserpine, whereas no change in enzyme activity followed administration of these drugs to 24 month old animals. In conclusion, increases in tyrosine hydroxylase activity in the brain that normally occur in the striatum of 3 month old rats after haloperidol or reserpine treatment are significantly decreased in 6 month old rats and not apparent in 24 month old rats.     

Effect of discontinuous short-wave electromagnetic field irradiation on the state of the endocrine glands

A study was made of the effect of interrupted EMF SW of 500 and 250 V/m on endocrine glands of male rats. The following changes were noted: activation of neurosecretion in hypothalamus, inhibition of adrenocorticotropic and gonadotropic functions of hypophysis and adrenal cortex, degenerative and dystrophic changes in testes. The observed disorders were aggravated as the intensity of the field and duration of exposure increased.     

Central catecholamine depletion inhibits peripheral lymphocyte responsiveness in spleen and blood

Experimental and clinical evidence has demonstrated extensive communication between the CNS and the immune system. To analyse the role of central catecholamines in modulating peripheral immune functions, we injected the neurotoxin 6-hydroxydopamine (6-OHDA) i.c.v. in rats. This treatment significantly reduced brain catecholamine content 2, 4 and 7 days after injection, and in the periphery splenic catecholamine levels were reduced 4 days after treatment. Central catecholamine depletion induced an inhibition of splenic and blood lymphocyte proliferation and splenic cytokine production and expression (interleukin-2 and interferon-gamma) 7 days after injection. In addition, central treatment with 6-OHDA reduced the percentage of spleen and peripheral blood natural killer (CD161 +) cells, and T-cytotoxic (CD8 +) cells in peripheral blood. The reduction in splenocyte proliferation was not associated with a glucocorticoid alteration but was completely abolished by prior peripheral sympathectomy. These data demonstrate a crucial role of central and peripheral catecholamines in modulating immune function.     

Effects of acute or chronic administration of desmethylimipramine on the local level of methionine incorporation in cerebral proteins in freely moving rats

Incorporation of methionine into brain proteins has been measured by quantitative autoradiography after desmethylimipramine administration in rats. Acute treatment decreased protein synthesis in three regions: hippocampus, hypothalamus and habenula. After chronic treatment the decrease extended to four other regions: substantia nigra, raphe dorsalis, trigeminal nerve and hypophysis. An opposite effect was observed in the retrosplenial cortex.     

Heavy load exercise induced dysfunction of immunity and neuroendocrine responses in rats

To determine whether immunity and neuroendocrine system is altered by different loads of exercise training in rats, eight-week-old male Sprague-Dawley rats were randomly assigned to one of the three groups: 1) cage control group (CCG); 2) moderate load training (MLT) (swimming at the intensity of 1.4 m/sec water flowing for 60 min per day); 3) heavy load training (HLT) (swimming at the intensity of 1.8 m/sec water flowing for 120 min per day). MLT and HLT rats were assigned to swim for 6 days per week for total of 6 weeks. All rats were sacrificed 36 h after their last training session. Splenocytes were pooled for assay of cell proliferation and neuropeptide contents in the hypothalamus, hypophysis and plasma were determined by radioimmunoassay while glucocorticoid specific binding in intact thymus was measured by radioligand binding assay. All rats were weighed weekly. The results showed that after 6-week training, rat splenocyte proliferation in response to Con A and LPS decreased in HLT rats compared with MLT and CCG rats. In addition, the contents of beta-endorphin, dynorphin A, arginine vasopressin and oxytocin in the hypothalamus, hypophysis and plasma were altered by HLT, as shown by increased plasma concentration of glucocorticoids and decreased glucocorticoids specific binding in intact thymus compared with MLT and CCG. Furthermore, a decreased body mass in HLT rats has been observed. The body mass of HLT rats was significantly lower than that in CCG and MLT rats at the end of the swimming training period. These data suggest that 6-week heavy load training induces the dysfunction of immunity and neuroendocrine responses, which might be one of the underlying mechanisms of immune dysfunction in overtraining.     

Technical note: removal of the unfragmented pituitary gland (hypophysectomy) in the rat.

A technique to excise the pituitary gland (hypophysis) in rats is described. The basisphenoid bone is reached from the ventral neck and is perforated to expose the pituitary gland and its stalk. An aspirator allows the removal of the hypophysis and the stalk, including pars tuberalis, in one piece. The advantages of this new technique include: 1) immediate verification of the entirety of hypophysectomy; 2) broad operating field which exposes the pituitary stalk up to the hypothalamus; 3) the use of tracheal intubation and artificial respiration to improve postoperative recovery and to allow expanded operation field even during prolonged surgery. Pre- and postoperative care are described. The mean survival rate after this type of operation was 79% in rats weighing 50 to 130 g and 90% in rats larger than 130 g.