Somatic afferent regulation of plasma luteinizing hormone and testosterone in anesthetized rats.

The effects of cutaneous stimulation on plasma luteinizing hormone (LH) and testosterone were examined in adult male Wistar rats anesthetized with pentobarbital. Under the resting condition without somatic sensory stimulation, plasma LH and testosterone measured every 30 min between 1400 and 1730 h revealed no significant fluctuations. Nociceptive mechanical stimulation of bilateral hindpaws by pinching for 10 min significantly increased plasma LH and testosterone. Plasma LH was increased 30 and 60 min after stimulation, while plasma testosterone was increased 60-150 min after stimulation. However, innocuous mechanical stimulation of bilateral hindlimbs by brushing for 10 min did not significantly change either plasma LH or testosterone. These results indicate that, when emotional factors were eliminated by anesthetization, cutaneous nociceptive information led to the increased secretion of LH from the anterior pituitary, resulting in an increase in testosterone secretion from the testes into the plasma.     

Differential plasma corticosterone responses to hippocampal stimulation

Summary The effect of limbic forebrain stimulation on pituitary-adrenal function was assessed by evaluating plasma corticosterone obtained prior to and following sham or electrical stimulation of urethane (1.20 g/kg) anesthetized female rats. Cortical electroencephalogram (EEG), electrocardiogram (ECG), heart rate (HR), mean arterial blood pressure (MAP), and respiration were routinely monitored. Timed blood samples (0.25 ml) were obtained from a catherized femoral artery. The HR (Bts/min), MAP (mm of Hg), and corticosterone levels (g/dl) for 7 non-stimulated rats averaged over 6 sampling periods were 385±19, 95±6, and 70.3±5.8 respectively. In electrically or sham stimulated rats, blood samples were taken just prior to stimulation (biphasic square waves, 100 A, 50 or 60 Hz, 1 ms, 1 s on/1 s off for 15 or 30 min) and 5, 10, 15, and 30 min after initiation of stimulation.Significant changes in plasma corticosterone levels were obtained following stimulation of hippocampal and amygdaloid areas. In contrast, no change in corticosterone concentration was observed following stimulation of cortex, corpus callosum, fornix and a variety of other CNS areas. Detailed analysis of hippocampal influence on urethane stimulated plasma corticosterone levels showed increased plasma corticosterone levels following stimulation of CA1. In contrast, stimulation of CA3, dentate (includes CA4) and the subiculum produced significant decreases in plasma corticosterone levels. No change in corticosterone levels was observed following sham stimulation. Collectively, these data indicate that consideration must be given to the possibility that differential neuroendocrine regulatory mechanisms reside within various limbic forebrain complexes and that electrical stimulation of limbic forebrain sites of urethane anesthetized rats may provide information regarding sites inhibitory to pituitary-adrenal activity.     

Hypothalamic corticotropin-releasing hormone (CRH) secretion into hypophysial portal blood is regulated by cutaneous sensory stimulation in anesthetized rats.

The effects of noxious and non-noxious mechanical stimulation of various segmental skin areas (face, forelimb and forepaw, abdomen, hindlimb and hindpaw) on the secretion of immunoreactive corticotropin-releasing hormone (iCRH) from the hypothalamus into hypophysial portal blood was examined in artificially ventilated rats under halothane anesthesia. Secretion of iCRH was calculated from the iCRH concentration in hypophysial portal plasma and the plasma flow rate. Noxious mechanical stimulation of the skin was delivered by pinching using surgical clamps, while non-noxious mechanical stimulation was provided by brushing with tooth brushes. Pinching of the bilateral forepaws or hindpaws and brushing of the bilateral hindlimbs for 20 min increased hypothalamic iCRH secretion. In contrast, pinching of the face or abdomen and brushing of the face, forelimbs, or abdomen for 20 min did not significantly influence it. These results indicate that cutaneous mechanical sensory stimulation contributes to the reflex regulation of CRH secretion from the hypothalamus into hypophysial portal blood, and also that this effect is highly dependent on the site of stimulation.     

Sympatho-adrenal hyperreactivity to footshock stress but not to cold exposure in spontaneously hypertensive rats

No significant differences in plasma levels of catecholamines were found in blood obtained via an indwelling tail artery catheter from awake, undisturbed spontaneously hypertensive (SHR) and normotensive Wistar-Kyoto (WKY) or NIH Sprague-Dawley rats. Although the blood pressures of SHR rats increased between 4 and 10 months of age, there were no changes in basal plasma catecholamine levels. Exposure to cold for 20 or 240 min resulted in significant increments in plasma catecholamine levels which were similar in SHR and WKY rats. In contrast, exposure to 5 min of inescapable footshock (2.5 mA, 0.4 sec duration, every 5 sec) produced striking increments in plasma catecholamines that were greater in SHR rats than in normotensive WKY or NIH Sprague-Dawley rats. SHR rats also exhibited greater levels of plasma corticosterone than WKY rats at 15 min after termination of footshock. Our results demonstrate that SHR rats are hyperadrenergic in their responses to some but not all forms of stressful stimulation.     

Glucocorticoid modulation of atrial natriuretic peptide, oxytocin, vasopressin and Fos expression in response to osmotic, angiotensinergic and cholinergic stimulation

The regulation of fluid and electrolyte homeostasis involves the participation of several neuropeptides and hormones that utilize hypothalamic cholinergic, alpha-adrenergic and angiotensinergic neurotransmitters and pathways. Additionally, it has been suggested that hypothalamus-pituitary-adrenal axis activity modulates hormonal responses to blood volume expansion. In the present study, we evaluated the effect of dexamethasone on atrial natriuretic peptide (ANP), oxytocin (OT) and vasopressin (AVP) responses to i.c.v. microinjections of 0.15 M and 0.30 M NaCl, angiotensin-II (ANG-II) and carbachol. We also evaluated the Fos protein immunoreactivity in the median preoptic (MnPO), paraventricular (PVN) and supraoptic (SON) nuclei. Male Wistar rats received an i.p. injection of dexamethasone (1 mg/kg) or vehicle (0.15 M NaCl) 2 h before the i.c.v. microinjections. Blood samples for plasma ANP, OT, AVP and corticosterone determinations were collected at 5 and 20 min after stimulus. Another set of rats was perfused 120 min after stimulation. A significant increase in plasma ANP, OT, AVP and corticosterone levels was observed at 5 and 20 min after each central stimulation compared with isotonic saline-injected group. Pre-treatment with dexamethasone decreased plasma corticosterone and OT levels, with no changes in the AVP secretion. On the other hand, dexamethasone induced a significant increase in plasma ANP levels. A significant increase in the number of Fos immunoreactive neurons was observed in the MnPO, PVN and SON after i.c.v. stimulations. Pre-treatment with dexamethasone induced a significant decrease in Fos immunoreactivity in these nuclei compared with the vehicle. These results indicate that central osmotic, cholinergic, and angiotensinergic stimuli activate MnPO, PVN and SON, with a subsequent OT, AVP, and ANP release. The present data also suggest that these responses are modulated by glucocorticoids.     

Acute increase in plasma corticosterone level in female mice evoked by pheromones

Exposure of a singly reared estrous female to grouped females or their bedding during 20 min resulted in an increase in the plasma corticosterone level during the first 10 min. The reaction of the adrenal glands was similar to the excitation evoked by stress connected with moving females from their own cage to a new, clean cage. The highest level of the adrenal hormone was present in estrous females after exposure to male bedding during 10 min. This was followed by a dramatic decrease during the next 90 min. It is suggested that male pheromones, through activation of the pituitary-adrenal system and acute release of corticosterone, participate in the stimulation of sexual behavior in receptive female mice.     

Modifications of adrenocortical responses following frontal cortex simulation in rats with hypothalamic deafferentations and medial forebrain bundle lesions

With the purpose of delineating the neural pathways in the rat which mediate adrenocortical responses following frontal cortex stimulation, the effects of partial hypothalamic deafferentations and medial forebrain bundle lesion were studied. In intact and sham-operated animals, cortical stimulation through permanently implanted electrodes caused a significant increase in plasma corticosterone levels. In rats with anterior hypothalamic deafferentation and bilateral medial forebrain bundle lesions the adrenal response to cortical stimulation was blocked completely, while in animals with posterior hypothalamic deafferentation there occurred a normal rise in plasma corticosterone. These studies demonstrate that the frontal cortex effects on adrenocortical secretion are neurally mediated and involve an anterior hypothalamic input, more specifically the medial forebrain bundle.     

Interaction of genotype and environment as determinants of corticosteroid response to stress

The temporal pattern of plasma corticosterone concentrations following electric shock was studied in 10-week old mice from two inbred strains (C57BL/10J and DBA/2) and their reciprocal crosses, half of which had been subjected to infantile stimulation. The roles of genotype, maternal factors, and handling were assessed as determinants of corticosteroid response to stress. Highly significant and opposing genetic and maternal effects interacted to limit extreme plasma corticosterone concentrations following stress. Infantile stimulation also acted to produce intermediate steroid responses.     

The preoptic area and bed nucleus of the stria terminalis are involved in the effects of the amygdala on adrenocortical secretion

In view of the role of the amygdala in the modulation of adrenocortical secretion we have studied the neural pathways which mediate this response. Changes in plasma corticosterone following medial amygdala stimulation, under pentobarbital anaesthesia, were studied in rats which chronically implanted electrodes in intact and lesioned animals. The rise in plasma corticosterone following amygdala stimulation was inhibited by bilateral lesions of the stria terminals, medial preoptic area, and bed nucleus of the stria terminalis, and to a greater extent by a combined lesion of the latter two structures. The combined lesion also completely blocked the adrenocortical response to olfactory stimulation. These various lesions did not affect, however, the rise in plasma corticosterone following ether stress. These data thus demonstrate that the stria terminalis, preoptic area and bed nucleus of the stria terminalis are involved in the transmission of neural impulses to the hypothalamus which activate adrenocortical secretion.     

The effects of infusions of synthetic adrenocorticotrophin in the conscious calf

1. A technique is described by which the whole of the effluent blood from the right adrenal gland can be collected as required from conscious, unrestrained calves. The technique may be used to measure adrenal blood flow gravimetrically and to compute the output of adrenal hormones under various conditions in the normal calf.2. In a group of seven calves mean cortisol output from the right adrenal gland was found to vary between 20 and 40 ng.kg(-1) min(-1) and corticosterone between 6 and 18 ng.kg(-1) min(-1) during a 2 hr period, 24 hr after surgery.3. Intravenous infusions of synthetic adrenocorticotrophin (5 ng.kg(-1) min(-1)) produced a significant increase in the output of both cortisol and corticosterone within 5 min. The output of both hormones rose to maximal values within 10-20 min and mean values of approximately 300 ng.kg(-1) min(-1) (cortisol) and 120 ng.kg(-1) min(-1) (corticosterone) were maintained thereafter for the duration of the infusion (120 min). The output of both steroids fell to values comparable with those observed initially within 45-60 min after the infusion was discontinued.4. These changes in glucocorticoid output in response to adrenocorticotrophin produced a significant rise in the concentration of both cortisol and corticosterone in peripheral plasma. It is noteworthy that the rise in the mean corticosterone concentration in the peripheral plasma was substantially less than that which might be expected from relating the rise in mean plasma cortisol concentration to cortisol output.5. The results of control experiments have eliminated the possibility that the sampling procedure might itself increase steroid output or peripheral plasma concentration. Comparison of results from calves of widely disparate ages (8-38 days) provided no evidence that either the resting output of cortisol or corticosterone or the response to adrenocorticotrophin changes with age within the range examined.6. Infusion of adrenocorticotrophin (5 ng.kg(-1) min(-1)) also stimulated an abrupt rise in adrenal blood flow; mean resting flow (210 +/- 23 mul.kg(-1)) increased by approximately 30% within 5 min and attained peak values (355-365 mul.kg(-1) min(-1)) between 10 and 30 min. Thereafter, adrenal blood flow steadily decreased and then fell rapidly to within the resting range when the infusion was terminated. No significant changes in heart rate or aortic blood pressure occurred during these infusions.7. The results are discussed in relation to those obtained in other species and under differing conditions by other workers.     

The effect of amygdaloid ablation or stimulation on plasma corticosterone levels in the male rat

Participation of the amygdaloid complex in pituitary-adrenal function was assessed by measuring plasma corticosterone levels after a variety of ablative or stimulatory procedures in male rats. Transection of the stria terminalis, ablation of the basolateral amygdalopiriform region or subtotal amygdalectomy did not alter a.m. or p.m. non-stress plasma corticosterone levels. Likewise, stress-induced increments in plasma corticosterone concentrations in experimental rats were not different from those in controls after 3 min exposure to ether vapor or immobilization in a supine position. As a corollary to the ablation studies, rats bearing chronically implanted electrodes were subjected to sham or electrical stimulation. In non-anesthetized (freely moving) rats electrical stimulation of the corticomedial amygdala, and basolateral amygdala, dorsal hippocampus and arcuate nucleus resulted in plasma corticosterone levels significantly (P < 0.05) greater than from those in sham stimulated controls. In contrast, in pentobarbital anesthetized rats plasma corticosterone concentrations in the amygdaloid and hippocampal stimulated animals were not different from those of sham stimulated rats; only stimulation of the arcuate nucleus produced plasma corticosterone levels greater (P < 0.05) than those in sham stimulated rats.These data indicate that in the male rat the amygdala is capable of influencing ACTH secretion. However, the finding that pituitary-adrenal function was not compromised by either selective or extensive amygdaloid ablation suggests that the amygdala is not essential for pituitary-adrenal function as we assessed this function.     

Electrical stimulation in the rat of the supraoptic nucleus: Failure to alter plasma corticosterone after surgical lesioning of the paraventricular nucleus

Electrical stimulation of the supraoptic nucleus (SON) for 3 min in dexamethasone-morphine-pentobarbital-pretreated rats raised the plasma level of corticosterone; 4 dats after placing a surgical lesion in the hypothalamic paraventricular nuclei (PVN) electrical stimulation of the SON failed to change the plasma level of corticosterone. We suggest that nerve fibres originating in or near the PVN that pass near the SON are likely to contain corticotropin releasing factor and elicit the release of ACTH following electrical stimulation.     

Plasma corticosterone fluctuations in an infant-learning paradigm

Plasma corticosterone fluctuations of infant rats were examined in a learning task. A blood sample for analysis of plasma corticosterone was collected from groups of 10- and 15-day-old pups following either no disturbance, standard maternal deprivation only, or deprivation and training in an established approach-mother, avoid-shock conflict task. In the latter condition, pups remained in the goalbox either alone or with an anesthetized dam for either 15 or 60 min before blood sample collection. Plasma corticosterone levels were elevated following deprivation plus training in pups of both ages and following deprivation only in the 15-day-old pups. Further, the presence of the dam in the goalbox reduced plasma corticosterone elevations, particularly among 15-day-old pups and at 60 min. These findings suggest that the mother's capacity to moderate the pup's plasma corticosterone response may contribute to her reinforcement value in infant-learning paradigms.     

Effects of olfactory bulbectomy and domicile on stress-induced corticosterone release in the rat.

Following bilateral olfactory bulbectomy or sham surgery, rats were housed either in groups of 5 or individually. After 10 days the animals were sacrificed and plasma corticosterone levels were measured. Half the animals in each group received predictable stress in the form of footshock immediately prior to exsanguination. Two levels of corticosterone elevation were noted, an intermediate level (circa 40 μg/100 ml plasma), in bulbectomised animals without stress and in sham operated animals with stress. Extreme corticosterone elevation (circa 80 μg/100 ml plasma) occurred in bulbectomised rats with stress. The type of housing had no effect on corticosterone elevation. The results are discussed in terms of a non olfactory function for the olfactory bulb, and the role of corticosterone and ACTH in acquisition learning.     

Locomotor activity,defecation score and corticosterone levels during an openfield exposure: A comparison among individually and group-housed rats,and genetically selected rat lines

Eight groups of rats, either differently reared or genetically selected, were compared in regard to locomotor activity, defecation score and plasma corticosterone levels during a 10 min openfield session. A relationship between defecation and corticosterone increases was found, whereas locomotion did not consistently correlate to the other parameters. The results were discussed in terms of emotionality, leading to the conclusion that isolation in rats reduces emotional responses in a novel stimulation.     

Reflex responses in plasma catecholamines caused by static contraction of skeletal muscle.

To examine a hypothesis of whether static muscle contraction produces a release of catecholamines from the adrenal medulla via reflex stimulation of preganglionic adrenal sympathetic nerve activity induced by receptors in the contracting muscle, we compared the reflex responses in a concentration of epinephrine (Ep) and norepinephrine (NEp) in arterial plasma during static contraction and during a mechanical stretch of the hindlimb triceps surae muscle in anesthetized cats. Static contraction was evoked by electrically stimulating the peripheral ends of the cut L(7) and S(1) ventral roots at 20 or 40 Hz. Mean arterial pressure (MAP) and heart rate (HR) increased 23 +/- 3.1 mmHg and 19 +/- 4.3 beats/min during static contraction. Ep in arterial plasma increased 0.18 +/- 0.072 ng/ml over the control of 0.14 +/- 0.051 ng/ml within 1 min from the onset of static contraction, and NEp increased 0.47 +/- 0.087 ng/ml over the control of 0.71 +/- 0.108 ng/ml. Following a neuromuscular blockade, although the same ventral root stimulation failed to produce the cardiovascular and plasma catecholamine responses, the mechanical stretch of the muscle increased MAP, HR, and plasma Ep, but not plasma NEp. With bilateral adrenalectomy, the baseline Ep became negligible (0.012 +/- 0.001 ng/ml) and the baseline NEp was lowered to 0.52 +/- 0.109 ng/ml. Neither static contraction nor mechanical stretch produced significant responses in plasma Ep and NEp following the adrenalectomy. These results suggest that static muscle contraction augments preganglionic adrenal sympathetic nerve activity, which in turn secretes epinephrine from the adrenal medulla into plasma. A muscle mechanoreflex from the contracting muscle may play a role in stimulation of the adrenal sympathetic nerve activity.     

Action of acth(1 -24) upon plasma corticosterone concentrations in racing pigeons (Columba Livia domestica)

To establish the optimal dose and sampling times for the ACTH stimulation test in racing pigeons, plasma corticosterone concentrations were measured in experimental animals at 0, 30, 60, 90, 120 and 150 min after intramuscular administration of different doses of ACTH(1-24). Baseline corticosterone concentrations varied between less than 0.2 and 1.24 mutg/dl. To evaluate adrenocortical function the clinician is advised to take a blood sample before and at 60 or 90 min after stimulation with 50 mug ACTH or at 30, 60, 90 or 120 min after stimulation with 125 mug ACTH. In healthy individuals a ten- to hundred-fold increase over baseline (t=0) corticosterone concentrations and absolute concentrations in the range of 2.2 to 15.0 mug/dl can be expected in the post stimulation sample.     

Sensitization of the plasma corticosterone response to novel environments

In Experiment 1, ICR Swiss mice were exposed to one of two novel cages for 1, 6, or 10 30-min trials. Plasma corticosterone elevations were greater in the more novel than in the less novel cage and following the sixth as compared to the first and tenth exposures. In Experiment 2, BALB/c and C57BL/6J mice and Sprague-Dawley rats also exhibited significant, though modest, increases in corticosterone levels from their first to their sixth 30-min exposures to the more novel cage. These findings demonstrate generalization of an apparent sensitization of the plasma corticosterone response across the two levels of novelty and four strains of laboratory rodents tested here. The transience of the effect may explain some earlier inconsistencies in the literature. In Experiment 3, five daily 30-min exposures to the more novel environment minimally raised resting corticosterone levels of ICR Swiss mice. Further, corticosterone levels during a first exposure were no greater at 15 or 45 min than at 30 min. These results suggest that the increase in corticosterone with repeated exposures reflects a true sensitization of the corticosterone response rather than an increase in resting levels with repeated exposures or a shift in the time course of the corticosterone response.     

Lack of effects of handling habituation and swimming stress on ethanol-induced motor impairment and GABAA receptor function.

The effects of handling habituation and swimming stress on ethanol-induced motor impairment and the GABAA receptor function were studied in adult male Wistar rats. Daily handling for 3 to 5 weeks had no significant effect on ethanol-induced motor impairment in the tilting plane test or on the activity of the rats in the elevated plus-maze test of anxiety. Plasma corticosterone concentrations were greatly elevated by the tilting plane test procedure, irrespective of handling habituation. Acute, 10-min swimming stress at +25 degrees C produced an elevated plasma corticosterone concentration comparable to that produced by the tilting plane test, again irrespective of handling habituation. In cerebrocortical homogenates, short-term swimming stress had no statistically significant effect on the muscimol stimulation of the GABAA receptor-mediated 36Cl- flux in handled and non-handled animals. Thus, handling habituation and stress had only minor effects on the activity of the central GABAergic systems in acute tests at behavioural and biochemical levels.     

Glucocorticoid receptor activation selectively hampers N-methyl-D-aspartate receptor dependent hippocampal synaptic plasticity in vitro

Corticosterone and exposure to stressful experiences have been reported to decrease hippocampal synaptic plasticity, in particular when relatively mild stimulation paradigms-presumably activating predominantly N-methyl-d-aspartate receptors-are being used. Using various stimulation paradigms and pharmacological approaches we tested therefore the hypothesis that elevated corticosterone levels, by activating glucocorticoid receptors, predominantly hamper N-methyl-D-aspartate receptor dependent synaptic plasticity in vitro. To address this, mouse hippocampal slices were treated for 20 min with corticosterone (100 nM) or vehicle and synaptic efficacy was examined 1-6 h later. First, we found that primed burst potentiation and synaptic potentiation after 10 Hz stimulation are predominantly N-methyl-D-aspartate receptor dependent, and are significantly suppressed after corticosterone treatment. Second, these latter effects were prevented by treating slices with the glucocorticoid receptor antagonist mifepristone prior to and during corticosterone administration. Third, theta burst potentiation, which was shown to involve activation of both N-methyl-D-aspartate receptors, voltage-dependent calcium channels and possibly other mechanisms, was not affected by corticosterone. However, theta-burst potentiation in the presence of nifedipine-singling out primarily the N-methyl-D-aspartate receptor dependent component-was reduced by corticosterone. These results indicate that corticosterone, via glucocorticoid receptor activation, selectively hampers N-methyl-D-aspartate receptor dependent synaptic plasticity in vitro and leaves more complex forms of long term potentiation unaffected. We speculate that these effects are involved in the impairment of cognitive performance by corticosteroid hormones after exposure to stressful and traumatic experiences.