Involvement of the sensory cortex in adrenocortical responses following photic and acoustic stimulation in the rat

In order to examine the possibility that the sensory cortex participates in the mediation of adrenocortical responses following sensory stimuli, the effects of photic and acoustic stimuli on plasma corticosterone were studied in rats with either visual or auditory cortex ablation. In animals with visual cortex ablation, the adrenocortical response to acoustic stimuli was intact; however, it was significantly reduced following photic stimulation. On the other hand, in animals with auditory cortex ablation, the response to acoustic stimulation was significantly reduced, but the response to photic stimulation remained intact. These data demonstrate the participation of the specific sensory cortex in adrenocortical responses following the stimulation of the corresponding sensory modality. The possible mechanisms involved may be either a tonic facilitatory effect of the specific cortex on subcortical mechanisms or the transmission of the specific stimuli in the primary sensory pathways, to achieve a full adrenocortical discharge.     

Norepinephrine depletion in the paraventricular nucleus inhibits the adrenocortical responses to neural stimuli

In view of the importance of teh paraventricular nuclei of the hypothalamus (PVN) in the regulation of adrenocortical secretion and the possible role of norepinephrine (NE) in this mechanism, we have studied the effects of injecting the catecholamine neurotoxin 6-hydroxydopamine into the PVN on basal corticosterone (CS) secretion and following ether stress and neural stimuli. PVN NE depletion did not affect the basal and the ether stress-induced rise in CS levels. However, it inhibited significantly the adrenal response to photic, acoustic and sciatic nerve stimulation, suggesting that the PVN NE plays a role in the activation of adrenocortical responses following afferent neural stimuli.     

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.     

Inhibition of adrenocortical responses following olfactory stimulation in rats with stria terminalis lesions

Previous experiments using hypothalamic deafferentation and brain lesions in rats have demonstrated that while anterior hypothalamic deafferentation blocked the adrenocortical response to olfactory stimulation completely, bilateral lesions in the ventolateral medial forebrain bundle and gemini region had only a partial effect. In order to elucidate the pathways mediating this response, the adreno-cortical responses to olfactory stimulation as well as to ether stress, photic, acoustic and sciatic stimulation were studied in rats with bilateral stria terminalis or small basolateral or corticomedial amygdaloid lesions. The adrenocortical responses to all the above modalities were normal and the only response which was completely inhibited was that to olfactory stimulation in rats with stria terminalis lesions.This would indicate that the stria terminalis serves as the hypothalamic afferent pathway activating the pituitary-adrenal reaction to olfactory stimulation.     

Effects of hypothalamic deafferentations on adrenocortical responses in the rat following hippocampal stimulation

Summary Adrenocortical responses, as expressed by changes in plasma corticosterone levels, following ether stress and dorsal hippocampal stimulation, were studied in intact rats and in rats with complete, anterior or posterior hypothalamic deafferentations. Ether stress produced normal responses in all experimental groups. In the three groups with hypothalamic deafferentations, the adrenocortical response following hippocampal stimulation was completely blocked, when compared to intact animals. The results suggest that the hippocampal signal enters the hypothalamus anteriorly, but that caudal propagation and posterior re-entry into the hypothalamus are also essential for the adrenocortical activation by the hippocampus.Supported by US-Israel Binational Foundation grant 1554/78     

Paraventricular nucleus neuronal responses following electrical stimulation of the midbrain dorsal raphe: evidence for cotransmission

Summary In order to determine the responses of paraventricular nucleus neurones following activation of central serotonergic pathways, single unit activity was recorded and responses following electrical stimulation of the midbrain dorsal raphe nucleus were examined. Excitation was recorded from approximately 50% of the cells, independent of whether they were antidromically identified as projecting to the median eminence or unidentified. Approximately 20% of cells were inhibited by the stimulation, the majority of these being unidentified. Parachlorophenylalanine-induced inhibition of serotonin synthesis reduced hypothalamic serotonin levels by 77% and caused a significant reduction in the proportion of cells excited by the stimulation, whereas the inhibitory responses were not affected. Intracerebroventricular administration of the serotonergic neurotoxin, 5,7-dihydroxytryptamine, which caused similar reductions in hypothalamic serotonin content (77%), reduced still further the proportion of excitatory responses and also reduced the proportion of cells inhibited by the stimulation. The data obtained suggest that serotonin acts as an excitatory neurotransmitter in the paraventricular nucleus; this is discussed particularly with respect to the regulation of the hypothalamo-hypophysial-adrenocortical axis. The loss of inhibitory responses in 5,7-dihydroxytryptamine treated, as opposed to the parachlorophenylalanine treated, animals suggests that the serotonergic fibers innervating the recorded cells may contain a cosecreted substance that may have important physiological actions in the control of neuronal activity in the region recorded.     

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.     

Serotonin regulation of corticoid secretion in infant rats

The plasma corticosterone response to various doses of serotonin and 5-hydroxytryptophan was studied in the 3-day-old rat. The maximum response to both drugs occurred 60 min after injection. Increased corticosterone concentration was observed at lower doses of serotonin than of 5-hydroxytryptophan. The effects of serotonin injection on Days 1–7 after birth on the plasma corticosterone response to 3 min of novelty stimulation at weaning was also studied. Serotonin treatment resulted in reduced adrenocortical reactivity which was reduced further when fluoxetine was given prior to the serotonin. Shocking rat pups on Days 1–7 also reduced adrenocortical reactivity and fluoxetine prior to shock further reduced adrenocortical reactivity when administered twice per day.     

Vascular endothelial growth factor-B is neuroprotective in an in vivo rat model of Parkinson's disease

Occlusal disharmony induced by placing an acryl cap on the lower incisors of rats is perceived as chronic stress. This chronic stress activates corticotropin-releasing hormone (CRH) neurons in the hypothalamic paraventricular nucleus (PVN), resulting in stimulation of the hypothalamic-pituitary-adrenal (HPA) axis. The ventral ascending noradrenergic bundles (V-NAB) from the brainstem innervate the PVN. To investigate the relationship between the response of the HPA axis and the V-NAB, we examined changes in extracellular noradrenaline (NA) in the PVN and plasma corticosterone, the final output of the HPA axis, following occlusal disharmony in rats injected with 6-hydroxydopamine (6-OHDA), a specific catecholamine neurotoxin. 6-OHDA microinjection into the V-NAB reduced the magnitude of the responses of extracellular NA in the PVN and the plasma corticosterone to occlusal disharmony. Our results suggest that V-NAB to the PVN are involved in occlusal disharmony-induced activation of the HPA axis.     

Body temperature and adrenal function in heat-exposed hypothalamic disconnected rats

Summary The effects of hypothalamic disconnection on body temperature and hypothalamo-pituitary-adrenal activity following acute and repeated exposures to heat were studied. Intact male rats, or animals with complete posterior or anterior hypothalamic disconnection, were exposed to a temperature of 36°C and a relative humidity of 35–45%. In the complete posterior and anterior hypothalamic disconnected rats the basalT _re was higher than that of the intact rats; the rise inT _re following heat exposure was lower in the operated rats than in the intact animals. All the experimental animals, except for those with anterior hypothalamic disconnection, showed a significant inhibition of corticosterone release on exposure to heat for 30 min, but no inhibition was observed in any of the disconnected rats when they were exposed to heat for 120 min. These results suggest that the main stimulus for ACTH release, during the first 30 min of heat exposure, is mediated by a neural input through the posterior hypothalamus and this is followed by a neural and/or humoral mechanism which enables the animals to increase their corticosterone secretion.     

Adrenocortical response following acute neurogenic stimuli is mediated by CRF-41

The present study examined whether neurogenic stimuli activate the pituitary-adrenal axis via CRF-41. Adult male rats were exposed to photic, acoustic or sciatic nerve stimulation. At 4, 15, and 30 min following the onset of stress, animals were sacrificed, trunk blood collected and the median eminence removed. At 4 min following the stress onset, there was a significant decrease in CRF-41 content of the median eminence, which persisted for 30 min. Concomitant with the decrease in CRF-41 content, serum adrenocorticotropic hormone (ACTH) and corticosterone levels increased. Thus, this study demonstrates that CRF-41 released from the median eminence plays a dynamic role in mediating the ACTH and corticosterone response to neurogenic stimuli.     

Discrimination and conditioning of photic evoked cortical macropotentials in the albino rat

Nine rats were rewarded with lateral hypothalamic brain stimulation for pressing a given bar following enhanced photic evoked components or for pressing another bar following depressed components. Evoking stimulus intensity was constant so that the responses to be discriminated varied due to presumably internal state fluctuations. Most rats learned only to generate one kind of wave and remain on the appropriate bar. Two rats that learned this operant learned additionally to signal trials in which they failed to generate the predominant response. One rat displayed discrimination in the absence of operant production of particular wave types. Specific electrophysiological correlates of these response styles were found and taken as evidence that operant neural conditioning may involve atypical states of the organism. Also discussed in terms of electrophysiological data were (1) the nature of the difficulty of neural discrimination, and (2) antecedents for the various observed response styles.     

Effects of individual housing on circadian rhythms of adult rats

Circadian rhythms of hypothalamic serotonin (5HT), its precursor tryptophan (TP) and its metabolite 5-hydroxy-indoleacetic acid (5HIAA), and of prolactin and corticosterone circulating hormones were determined in group-housed and in individually-housed male rats, adapted to a 12:12 light/dark cycle. After 5 weeks of individual housing, 5HT peaked later, TP and 5HIAA peaked earlier, and the mesor level of TP and 5HIAA decreased with respect to group-housed animals. Individual housing caused an increase in the corticosterone mesor level, but did not affect amplitude or acrophase. The circadian rhythm of prolactin was unchanged by individual housing.     

Osteopontin stimulates invasion of NCI‐h295 cells but is not associated with survival in adrenocortical carcinoma

Gene array studies indicated that osteopontin (OPN) mRNA is highly expressed in adrenocortical carcinomas (ACCs). OPN enhances invasiveness, proliferation, and metastasis formation, and is associated with poor survival in some malignant diseases. Integrin αvβ3 has been shown to mediate OPN effects on invasion. In this study, we demonstrated OPN and integrin αvβ3 expression in normal adrenal glands and benign adenomas, with staining seen exclusively in adrenocortical cells as well as even stronger staining in ACC. Western blot analysis confirmed overexpression of OPN in ACC (p < 0.01). With Matrigel invasion assays, we have shown that OPN greatly stimulates the invasiveness of NCI‐h295 cells (>six‐fold increase, p < 0.001). Transfection with integrin αvβ3 further increased invasiveness after OPN stimulation (p < 0.001). This increase was reversed by the addition of an anti‐integrin β3 antibody, indicating a functional relationship of OPN and integrin αvβ3 in ACC. With tissue arrays, we confirmed high OPN expression in 147 ACC samples. However, no association with survival was seen in Kaplan‐Meier analysis including 111 patients with primary tumours graded for OPN staining and follow‐up data available. In conclusion, our in vitro data indicate that OPN and integrin αvβ3 may act as a functional complex facilitating the invasiveness of adrenocortical tumours. This relationship remains of relevance to our understanding of carcinogenesis, but further studies are needed to address the physiological and pathological function of OPN in adrenal tissue. Copyright © 2009 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.     

Neuropeptides TRH and cyclo(His-Pro) share neuromodulatory,but not stimulatory,action on hypothalamic neurons in vitro: implication for the regulation of feeding

Summary Electrical activity of neurons in rat hypothalamic ventromedial nucleus was recorded in tissue slices, to investigate central neural mechanisms underlying reduction of food intake caused by TRH and its metabolite, cyclo(His-Pro) [cHP]. Application of TRH had two actions: stimulation of neuronal activity, which was desensitized on closely repeated applications; and modulation of neuronal responses to neurotransmitters, even in the absence of the stimulatory action. The neuromodulatory but not the direct stimulatory action could also be achieved by cHP. The neuromodulatory action is more likely to be a neural mechanism underlying the inhibition of feeding, while other biological functions, unique to TRH, may depend on direct stimulation. In this way, TRH could achieve different biological results through different modes of action on hypothalamic neurons.     

Paraventricular nucleus magnocellular neuronal responses following electrical stimulation of the midbrain dorsal raphe

Summary In order to determine the responses of paraventricular nucleus magnocellular neurones following activation of central serotonergic pathways, single unit activity was recorded and responses following electrical stimulation of the midbrain dorsal raphe nucleus were examined. Approximately one third (32%) of the phasically active, vasopressin-secreting neurones were inhibited by the stimulation, the remaining such cells being nonresponsive. In contrast, only two of the non-phasic cells (13%) were inhibited by the stimulation whilst 53% were excited (p< 0.005, chi²-test). The onset latency of both inhibitory and excitatory responses were similar, whilst offset of the inhibitory responses was about twice that of the excitatory responses (p < 0.005, t-test). Two of the nonphasic cells were antidromically identified as projecting to the dorsal raphe. The results obtained indicate a role for dorsal raphe projections to the paraventricular nucleus in the regulation of neurohypophysial hormone secretion. The observation that different sub-populations of the cells recorded showed different responses, suggests that several mechanisms may be involved in the control of neuronal activity in the region recorded, in response to activation of the central serotonergic pathway examined. The results obtained are intended to further clarify the neural mechanisms regulating the secretion of vasopressin and oxytocin from the neurohypophysis.     

Circadian rhythms of hypothalamic norepinephrine and of some circulating substances in individually housed adult rats.

The circadian rhythms of hypothalamic norepinephrine (NE) and of circulating norepinephrine, epinephrine (E), corticosterone, aldosterone, and serotonin (5HT) were determined in group-housed and in individually housed male adult rats, adapted to a 12/12 light/dark cycle. After 5 weeks of individual housing, compared to group-housed animals, hypothalamic NE mesor decreased, while circulating NE, E, and corticosterone mesor increased. The circadian rhythms of aldosterone and 5HT were unaffected by individual housing.     

Glucocorticoid hypersecretion following intracerebroventricular injection of ethylcholine mustard aziridinium ion in rats

To investigate whether cholinergic hypofunctions in the brain influence hypothalamic-pituitary-adrenal activity, we examined the effects of cholinergic neurotoxin ethylcholine mustard aziridinium ion on basal and stress-induced levels of corticosterone in rats. Blood sampling from rats following intracerebroventricular injection of saline (5 microl, as a control) or this neurotoxin (5 nmol/5 microl) was performed over a day in one series, and was taken before, during and after an immobilization stress exposure in another series. Plasma levels of corticosterone and adrenocorticotropin were determined by the radioimmunoassay. The basal levels of plasma corticosterone and adrenocorticotropin over a day were significantly higher in the neurotoxin-treated rats, compared with the control rats (corticosterone, P<0.001; adrenocorticotropin, P<0.05). Further, relative adrenal gland weight of the neurotoxin-treated rats was significantly greater than that of the control rats (P<0.05). However, responses in plasma corticosterone level caused by the immobilization stress in the neurotoxin-treated rats were not different from those in the control rats.The present study demonstrated that damage to the cholinergic neurons in the brain increased hypothalamic-pituitary-adrenal activity over a day, probably due to freedom from inhibitory influences of the hippocampal cholinergic system, but that this damage did not influence stress-induced changes in plasma glucocorticoid level.     

Plasma corticosterone response to serotonin altering drugs in the 3-day-old rat.

Three-day-old rats were injected with various neurotransmitter altering agents to demonstrate a functional relationship between these drugs and plasma corticoid levels. Plasma corticosterone levels were increased after injection of methiothepin, methysergide, and 5-hydroxytryptophan (5-HTP), but were not changed by cholinergic, adrenergic, and dopaminergic compounds. Imipramine alone had no effects on plasma corticoids but in combination with 5-HTP resulted in a more sustained response than 5-HTP alone. Afunctional relationship between plasma corticosterone and serotonin receptors has been demonstrated in the 3-day-old rat. The presence of this relationship just after birth suggests the possibility that serotonin may be a mediator of early experience effects on later adrenocortical function.     

Pathologic features and expression of insulin-like growth factor-2 in adrenocortical neoplasms

We analyzed a series of adrenocortical neoplasms to compare the clinicopathologic features and the expression of insulin-like growth factor-2 (IGF-2) in adrenocortical adenomas and carcinomas. IGF-2 is a growth factor commonly expressed in many tumors including adrenal cortical and medullary neoplasms. Formalin-fixed paraffin-embedded tissues from 64 adrenocortical adenomas and 67 adrenocortical carcinomas were analyzed. The carcinomas were histologically graded from 1 to 4 based on mitotic activity and necrosis. Tumor weight, size, and follow-up information were obtained by chart review. Expression of IGF-2 was detected by immunohistochemistry with the avidin-biotin-peroxidase complex method and a monoclonal antibody against IGF-2. Adrenocortical carcinomas were larger (mean: 13.1 cm, 787 g) than adenomas (mean: 4.2 cm, 52 g) (p<0.001). In patients with adrenocortical carcinomas, high tumor grade (3 or 4) (p=0.01) was associated with decreased survival. Expression of IGF-2 was higher in adrenocortical carcinomas than in adenomas (p<0.001). These results show that tumor size and weight along with expression of IGF-2 protein are useful features to assist in distinguishing between adrenocortical adenomas and carcinomas, and that high tumor grade is a predictor of survival in adrenocortical carcinomas. However, single immunohistochemical markers such as IGF-2 or single histopathologic features cannot by themselves separate adrenocortical adenomas from carcinomas, and a combination of clinical, gross, and microscopic features are needed to establish the diagnosis in difficult cases.