Changes in the leukocyte distribution and surface expression of adhesion molecules induced by hypothalamic stimulation in the cat.

Emotions and the neuroendocrine system are known to affect leukocyte distribution. However, there have so far been few reports on the relationship between hypothalamically induced emotional behavior and the endocrine-immune response. We previously reported changes in the leukocyte distribution and adhesion molecules induced by anteromedial hypothalamus stimulation (AH stimulation), which elicits restlessness behaviors in the cat. In this study, we examined ventromedial hypothalamus stimulation (VMH stimulation), which elicits threat behaviors. In addition, the endocrine responses after VMH stimulation were evaluated. VMH stimulation as well as AH stimulation induced elevations of plasma cortisol and epinephrine levels and granulocytosis and lymphopenia. In contrast, VMH stimulation induced only an elevation of plasma norepinephrine and elicited an opposite pattern of CD62L expression on the leukocyte subpopulations. The different endocrine-immunological reactions between VMH stimulation and AH stimulation were thus associated with different types of behavioral responses.

Hypothalamically induced emotional behavior and immunological changes in the cat.

Numerous animal studies on the correlation between stress and immunity have been performed but few such studies have been made concerning the relationship between various kinds of stress-related emotional behavior and immunological changes. Electrical stimulation of the hypothalamus in cats elicits various emotional behaviors such as restlessness, defensive attack, defensive retreat and quiet biting attack. We examined changes in the lymphocyte proliferative responses and plasma cortisol level which accompanied such emotional behavior. A significant increase in plasma cortisol was observed in the restlessness, defensive attack and defensive retreat groups, but not in the quiet biting attack or non-response (control) groups. A significant increase in the lymphocyte proliferative responses to phytohemagglutinin (PHA) was observed in the restlessness and defensive attack groups but not in the defensive retreat, quiet biting attack or non-response groups. These results suggest that various kinds of emotional behavior appear to be differentially correlated with the lymphocyte proliferative responses, while also being differentially correlated with the plasma cortisol concentration. Because the changes in lymphocyte responses and plasma cortisol did not always completely correlate with one another, the changes in the lymphocyte responses are not considered to be influenced by plasma cortisol alone.

Changes in the leukocyte distribution and surface expression of adhesion molecules accompanied with hypothalamically induced restlessness in the cat.

One type of emotional behavior called restlessness occurs when the anteromedial hypothalamus is stimulated in cats. We examined the changes in the distribution and surface expression of adhesion molecules in leukocytes accompanied with restlessness. Mature female cats were used for this study. The cats were stimulated with 60 Hz sine wave train pulses (20-90 microA, 10 s in duration, at 5-min intervals) for 60 min. Samples of blood were collected from 30 min before stimulation up to several hours after the final stimulation. The number of granulocytes increased just after stimulation, while at the same time the expression of L-selectin decreased. On the other hand, the number of CD4+ and CD8+ T lymphocytes decreased at 1-2 h after the end of the stimulation, while the expression of L-selectin increased. In addition, the expression of LFA-1 and VLA-4 did not change. These data suggest that hypothalamically elicited restlessness is thus accompanied by a leukocyte distribution change, which might be mediated by changes in the expression of L-selectin on leukocytes. Plasma cortisol increased during stimulation in restlessness. However, during in vitro culture experiments, cortisol did not alter the expression of leukocyte L-selectin which thus indicated that cortisol does not directly affect the surface expression of L-selectin. These results thus suggest that hypothalamically induced restlessness is a useful stress model for psychoneuroimmunological studies.

The effect of interstimulus intervals and between-block rests on the auditory evoked potential and magnetic field: is the auditory P50 in humans an overlapping potential?

OBJECTIVE: The human auditory P50 may consist of overlapping potentials. To test this hypothesis, we manipulated interstimulus intervals (ISIs) and between-block rests, and recorded the P50, P50m, N100 and the N100m simultaneously. METHODS: Subjects were 12 right-handed healthy adults. Four conditions included: (1) 1.5 s/rest, (2) 1.5 s/successive, (3) 0.5 s/rest, and (4) 0.5 s/successive. Auditory stimuli were presented a total of 880 times for each condition. Auditory evoked potentials and magnetic fields were recorded. We examined the P50, N100, P50m, N100m and dipoles of the P50m and the N100m. RESULTS: There was no significant effect of the ISI on the P50 amplitudes, but the P50m amplitudes in the 0.5 s ISI conditions were significantly smaller than those in the 1.5 s ISI conditions. The N100 and the N100m amplitudes in the 0.5 s ISI conditions were significantly smaller than those in the 1.5 s ISI conditions. The N100 and the N100m amplitudes in the resting conditions were significantly larger than those in the successive conditions. The P50m dipoles were slightly deeper and more anterior than those of the N100m in primary auditory cortex. CONCLUSIONS: Central structures other than supratemporal cortex contribute to the P50 and that the P50 in humans represents overlapping potentials.

Effects of both the emotional behavior and feeding conditions on the circulating plasma volume and plasma glucose levels in cats.

Influence of hypothalamically induced emotional behavior on the circulating plasma volume, plasma levels of glucose, epinephrine (E), norepinephrine (NE), dopamine (DA) and cortisol were examined in awake cats under both fasted and fed conditions. Restlessness was evoked intermittently for 6 h by electrical stimulation of the anteromedial hypothalamus (AMH). Blood was sampled immediately before, 1 h after and 6 h after the start of stimulation. Changes in the plasma volume was calculated by changes of hemoglobin (Hb) and hematocrit (Ht). As the control group, another 7 cats with electrodes implanted but unstimulated were identically treated under both fasted and fed conditions. Both E and glucose levels in restlessness group once markedly increased after 1 h and then tended to decrease after 6 h, whereas NE levels in restlessness group increased after 1 h and further increased after 6 h, whether cats were fasted or fed. DA levels increased under the fasted condition of restlessness. The cortisol level markedly increased in both fasted and fed restlessness groups. The plasma volume in control group increased under the fed condition, while in restlessness group it decreased remarkably and tended to decrease more in a fasted state than in a fed state. These results indicated that AMH induced restlessness elicited marked sympatho-adrenal activation, hyperglycemia and hemoconcentration, whether cats were fasted or fed. Relationship among such responses, and the difference in responses between fasted and fed conditions were also discussed in the paper.

Ceruletide inhibits phencyclidine-induced dopamine and serotonin release in rat prefrontal cortex.

Phencyclidine (PCP; 5.0 mg/kg, i.p.) produced a greater increase in extracellular dopamine (DA) levels in the prefrontal cortex than in the striatum, while PCP increased the extracellular 5-hydroxytryptamine (serotonin; 5-HT) levels in the prefrontal cortex but not the striatum, as determined by in vivo microdialysis in awake, freely moving rats. The cholecystokinin (CCK)-related decapeptide ceruletide (120 and 400 microg/kg, i.p.), administered 60 min prior to PCP, significantly attenuated the PCP-induced increase in the extracellular levels of DA and 5-HT in the prefrontal cortex, but not in the striatum. These effects were reversed by PD 135,158, a selective CCK-B receptor antagonist (0.1 mg/kg, s.c.), administered 5 min prior to ceruletide. When administered alone, ceruletide (400 microg/kg, i.p.) significantly increased basal extracellular DA levels only in the prefrontal cortex. The selective N-methyl-D-aspartate (NMDA) receptor antagonist dizocilpine (0.5 mg/kg, i.p.) also increased extracellular DA levels in the prefrontal cortex, but this effect was unaffected by ceruletide pretreatment. These results suggest that ceruletide may differentially modulate basal and PCP-induced release of DA and 5-HT in the prefrontal cortex.

IL-1beta increases norepinephrine level in rat frontal cortex: involvement of prostanoids, NO, and glutamate.

The effects of local administration of interleukin-1beta (IL-1beta) were studied by using an intracerebral microdialysis technique in rats. A local injection of IL-1beta (3 and 10 ng) induced an elevation of norepinephrine (NE) concentration in the medial prefrontal cortex (mPFC). IL-1-receptor antagonist (800 ng) completely blocked the IL-1beta-induced NE increase. Diclofenac, a cyclooxygenase inhibitor (500 microM), and Nomega-nitro-L-arginine, a nitric oxide (NO) synthase inhibitor (100 microM), applied through the dialysis probe, did not affect the initial rise in NE levels observed 20 min after injection of IL-1beta but completely suppressed the late phase of IL-1beta-induced NE increase at 40 min and thereafter. In contrast, local perfusion of 6-cyno-7-nitroquinoxaline-2,3-dione, a non-N-methyl-D-aspartic acid (NMDA) glutamate-receptor antagonist (50 microM), but not DL-2-amino-5-phosphonovaleric acid, an NMDA-receptor antagonist (100 microM), blocked both phases of IL-1beta-induced NE increase. Furthermore, a microinjection of IL-1beta elevated the extracellular concentration of glutamate in the mPFC. These findings suggest that the IL-1beta-induced rise in NE levels in the mPFC is caused by activation of the glutamatergic system and the glutamate-induced increases in prostanoids and NO.

Distribution and co-localization of nitric oxide synthase and argininosuccinate synthetase in the cat hypothalamus.

Argininosuccinate synthetase (ASS) and nitric oxide synthase (NOS) comprise part of the cyclic metabolic pathway to produce nitric oxide (NO). ASS is one of the arginine synthesis enzymes which synthesizes argininosuccinate from aspartate and citrulline, and NOS forms NO and citrulline from arginine. This study examines the localization of ASS and NOS in the cat hypothalamus using nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d) histochemistry and immunohistochemistry against ASS and NOS. NADPH-d positive and/or ASS-immunoreactive neurons were localized in the following areas: the anterior hypothalamic area, the anterior hypothalamic nucleus, the supraoptic nucleus, the suprachiasmatic nucleus, the periventricular complex, the paraventricular nucleus, the parvocellular nucleus, the lateral hypothalamic area, the dorsomedial hypothalamic nucleus, the dorsal hypothalamic area, the posterior hypothalamic area, and the supramammillary nucleus. NOS and ASS double-labeled neurons were found in the anterior hypothalamic area, the supraoptic nucleus, the central part of the paraventricular nucleus of the hypothalamus, the lateral hypothalamic area, ventral part of the parvocellular hypothalamic nucleus, the posterior hypothalamic area, and the supramammillary nucleus. Double-labeled neurons in the hypothalamus comprised 20.7-32.0% of ASS-immunoreactive neurons and 10.2-26.3% of NOS-immunoreactive neurons. The results suggest the existence of the 'NO cycle' in situ and the physiological importance of NO and argininosuccinate in several regions of the cat hypothalamus.

Relationship of emotional behaviors induced by electrical stimulation of the hypothalamus to changes in EKG, heart, stomach, adrenal glands, and thymus.

The relationships of hypothalamically elicited emotional behaviors to their accompanying pathophysiological effects were examined as a model of how complex "emotional behaviors" may be related to fundamental psychosomatic disorders. Twenty-two unanesthetized adult cats were studied. EKG alterations and histological changes in the heart, stomach, adrenal glands, and thymus were related to the specific stereotypical emotional behaviors that could be elicited by hypothalamic stimulation in tamed subjects. Restlessness, threat, and searching-biting behaviors were evoked by electrical stimulation of the anteromedial, ventromedial, and lateral hypothalamus, respectively. The occurrence of cardiac arrhythmias, ST and/or T (ST-T) changes in the EKG, histological damage to myocardium, gastric erosion, and adrenal hyperplasia were generally observed in the restlessness and threat groups but not in the searching-biting group. The pathophysiological effects were similar in the restlessness and threat groups with no specific EKG change or organ effect attributable to either site of stimulation. Hypothalamically elicited restlessness or threat behaviors in cats are each associated with cardiac, gastric, and adrenal pathophysiologies.

The relation of emotional behavior to plasma catecholamines, cortisol and ventricular arrhythmia.

Hypothalamic stimulation applied through chronically implanted electrodes elicits several kinds of emotional behavior in conscious cats. We chose 3 kinds of emotional behavior, i.e., restlessness, threat and searching-biting. Under lightly anesthetized condition, we examined the changes of E, NE, DA and cortisol levels in arterial plasma and the cardiovascular responses (changes of systolic and diastolic blood pressure, heart rate and the occurrence of poststimulus ventricular arrhythmia) associated with electrical stimulation of specific sites within the hypothalamus. Both in restlessness and threat groups, elevation in systolic blood pressure was significantly greater and also the occurrence of ventricular arrhythmia and elevation in diastolic blood pressure tended to be greater than in the searching-biting group. Plasma E, NE, DA and cortisol increased significantly in restlessness and threat groups but remained unchanged in searching-biting and control groups. The ratios of changed values in catecholamines: delta E/delta NE, delta E/delta DA or delta NE/delta DA were not significantly different between any groups of emotional behavior. Furthermore, in the restlessness group, delta E, delta NE and delta DA all showed significant correlation with both the number of ventricular arrhythmia and changes of diastolic blood pressure; and the number of ventricular arrhythmia showed significant correlation with both the changes of diastolic blood pressure and heart rate. None of these correlations was observed in the threat or searching-biting group. These results suggested that restlessness and threat behaviors were more closely related to stress response than searching-biting behavior in cats. The differences in the endocrine and cardiovascular responses between restlessness and threat behavior were also discussed in the paper.

Relation of emotional behaviors to urine catecholamines and cortisol.

We examined changes of epinephrine, norepinephrine, and cortisol levels in 24-h urine accompanying emotional behavior in cats such as restlessness, threat, and quiet biting attack elicited by electrical stimulation of specific sites within the hypothalamus. Although norepinephrine remained unchanged with restlessness but increased with threat, elevation of epinephrine and cortisol levels was common to restlessness and threat. No significant changes in these hormonal levels were seen with quiet biting attack and control. Therefore, it was suggested that emotional behaviors such as restlessness and threat are more closely related to emotional stress than quiet biting attack in cats.

Hypothalamic linkage in stress-induced hypocalcemia, gastric damage, and emotional behavior in rats.

The effects of hypothalamic lesions on stress-induced hypocalcemia, gastric damage, and swim test-evoked behavior were examined in rats. Bilateral lesions of the ventromedial nucleus in the hypothalamus (VMH) eliminated water-restraint stress-induced hypocalcemia and attenuated any gastric damage compared with those in the sham-operated rats. In contrast, lesions in the paraventricular nucleus (PVN) exacerbated both the stress-induced hypocalcemia and gastric lesions in comparison with those in the control rats. In a forced-swimming test, the VMH-lesioned rats showed a significantly shorter time of immobility as well as a longer duration of struggling than the control rats, respectively, while the PVN-lesioned animals spent a longer time in immobility and a shorter period struggling than the control rats. These results suggest that the VMH has an accelerative action in stress-induced hypocalcemia, gastric lesions, and behavioral despair, while the PVN has an opposite effect.

Effect of dietary vitamin E on lipofuscin accumulation with age in the rat brain.

We investigated the effect of dietary vitamin E on lipofuscin accumulation with age in the hippocampus, the inferior olive and the cerebellum of young (3-5 months old) middle-aged (12-14 months old) and old (24-26 months old) male Sprague-Dawley rats. The rats were fed either a vitamin E-deficient diet, vitamin E-supplemented diet or a control diet after reaching four weeks old. We employed both quantitative light microscopy using semithin sections and qualitative fluorescence microscopy for the analysis of lipofuscin accumulation with age. The concentrations of alpha-tocopherol were measured simultaneously in both the plasma and the three brain regions investigated. The effect of vitamin E deficiency was statistically significant only in the inferior olive of young rats and in all the three brain regions of middle-aged rats. The effect of vitamin E supplementation was statistically significant in all three brain regions of middle-aged rats. There was no statistically significant effect of vitamin E deficiency or supplementation on lipofuscin accumulation with age as compared with the control rats in all three brain regions of old rats. It was thus revealed that dietary vitamin E clearly had a significant effect on lipofuscin accumulation with age in the rat brain up until middle age, and that the same effect became indistinct in the latter half of their life.

Effect of fish oil and safflower oil in common Japanese diet on human plasma fatty acid composition.

The influence of fish oil and safflower oil contained in the common Japanese diet as the main dietary polyunsaturated fatty acid source on plasma fatty acids in ten female student volunteers (21-22 years old) was investigated. The subjects were divided into two groups and fed the experimental diets for five days. The total daily fat intake in the fish diet and safflower oil diet was 54.4 g and 56.2 g, respectively, and the fat derived from fish and safflower oil was 16 g and 23 g, respectively. The proportion of linoleic acid was reduced in the plasma of subjects fed the fish diet and increased in the plasma of subjects fed the safflower oil diet. The plasma levels of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) were significantly elevated in the fish diet group. The ratio of EPA/arachidonic acid (AA) was higher, and those of n-6/n-3 and n-9/n-3 were lower in the plasma of subjects fed the fish diet when compared to the results obtained from plasma of subjects fed the safflower oil diet. From these results, it seems likely that fish oil in the common Japanese diet is a favorable source of plasma EPA and DHA even in such short term supplementation and with such a small amount of daily consumption.

Inhibitory effects of acetylcholine on aggressive-defense reaction induced by electrical stimulation of the hypothalamus in cats.

The effects of ventral amygdalofugal pathways (VAF) or stria terminalis (ST) on the hypothalamic aggressive-defense (rage) reaction were examined in fifteen cats. After bilateral electrolytic lesions of the VAF or ST, 0.5 pmol of acetylcholine (ACh) was injected into the VMH and electrical stimulation was applied. The changes in the threshold for range reaction were compared in the non-lesioned, VAF-lesioned and ST-lesioned group. Both VAF-lesioned and non-lesioned cats showed a similar degree of significant elevation in the threshold for rage at 5 min after injection, while ST-lesioned group had no significant elevation. These results suggest that the ST is concerned with the elevation of threshold for rage due to ACh injection.

Emotional behavior and arrhythmias induced in cats by hypothalamic stimulation.

As the relationship between emotional behavior and electrocardiographic (ECG) change induced by hypothalamic stimulation is poorly understood, eighty-four points in various areas within the hypothalamus in conscious cats were stimulated electrically through chronically implanted electrodes, the objective being to clarify the behavior accompanying ECG changes, in particular poststimulus arrhythmias. Forty-one of 84 points elicited behavioral patterns such as defense reaction, pseudo-rage and restlessness (classified as group A), and in twenty-one (51%) of these 41 points arrhythmias occurred after cessation of stimulation. Forty-three of 84 points elicited behavioral patterns including predatory, exploratory and other behavioral responses (classified as group B), and in three (7%) of 43 points, poststimulus arrhythmias followed. Under light anesthesia, stimulations of twofold current intensity were applied at these points, and the incidences of the arrhythmias did not change in either group. The arrhythmia-inducing area in the cases of group A was found to lie dorsal and caudal to the optic chiasma and to extend caudally in the fornix. Three points in the cases of group B were located in the outer area of the aforementioned area. These studies showed that arrhythmias and group A behavior were observed mainly from stimulation of the anterior hypothalamus, whereas stimulation of other areas of the hypothalamus, including the lateral and the posterolateral hypothalamus, produced group B behavior and no arrhythmias.

Fine structures of the cerebrospinal fluid-contacting neurons in the hypothalamus of the lamprey, Lampetra japonica.

In the preoptic, infundibular and posterior recesses of the lamprey, Lampetra japonica cerebrospinal fluid (CSF)-contacting neurons are distributed much more abundantly than in higher vertebrates. They are classified into three different types on the basis of their granules size or electron density: type 1 containing round dense granules of large size (180--230 nm in diameter), type 2 containing variously shaped dense granules of medium size (150--200 nm) and type 3 containing variously shaped granules of small size (100--150 nm). The neurons of the third type may be further classified into two subtypes: type 3a possessing dense granules and type 3b showing less dense granules. The intraventricular process of CSF-contacting neurons contains Golgi cisternae that are forming new secretory granules and well developed granular endoplasmic reticulum with dilated cisternae. It is suggested that secretory granules might be formed in the intraventricular process in addition to the perikaryon. Besides the small cytoplasmic protrusions presumably involved in microapocrine secretion, intraventricular processes occupied with many empty granules are detected which might represent a process of diacrine secretion. The intraventricular processes further contain multivesicular bodies which incorporate old or overproduced secretory granules, suggesting a process (crinophagy) and large lipid droplets likely corresponding to the final digestive product. The CSF-contacting neurons of all types are usually supplied with axo-somatic synapses on the perikaryon and subsurface cisternae are sometimes observed beneath the postsynaptic membrane. Axon terminals are also found forming synapses on the intraventricular process of type 2 and 3 neurons.