Kunio Kitahama

Demonstration of GABAergic cell bodies in the suprachiasmatic nucleus: In situ hybridization of glutamic acid decarboxylase (GAD) mRNA and immunocytochemistry of GAD and GABA

The existence of GABAergic neurons in the rat suprachiasmatic nucleus (SCN) was demonstrated by three specific markers; mRNA coding for glutamic acid decarboxylase (GAD) and visualized by in situ hybridization using a 35S-labelled cDNA probe, and GAD protein and GABA were identified by immunocytochemistry using specific antisera. In situ hybridization demonstrated well labelled GAD mRNA positive cells throughout SCN, and GABA and GAD immunoreactive cells showed similar distributions. These results indicate that GABA is a transmitter of a large portion of the SCN neuronal population.

Increased paradoxical sleep in mice during acquisition of a shock avoidance task.

Abstract Two strains of mice were subjected to a complex shock avoidance task. The C57BR (brown) strain were superior to the C57BL (black) strain in their learning ability. Both strains showed a long term increase in paradoxical sleep (PS) prior to the maximum increase in learning performance (MIP). For the brown starin, this increase was apparent in the 24 h before the MIP. For the black strain, the increase began 48 h prior to the MIP. In the brown strain the day after the MIP, a second effect appeared. PS was higher in the first half hour after sleep onset, following the training session. This increase was due to the larger numbers of slow wave sleep-paradoxical sleep (SWS-PS) cycles. The effect was not present in the black strain. It was concluded that two mechanisms are probably at work during the learning process, one during the earlier stages and one during the later stages as learning reaches criterion.

Dopamine- and DOPA-immunoreactive neurons in the cat forebrain with reference to tyrosine hydroxylase-immunohistochemistry

The distribution of cell bodies containing immunoreactivities to dopamine (DA), L-3,4-dihydroxyphenylalanine (DOPA) and tyrosine hydroxylase (TH) was studied immunohistochemically in the cat forebrain especially in the hypothalamus with or without intraventricular administration of colchicine. In normal cats, DA-immunoreactive (IR) neurons, whose intensity of immunostainings was variable from one to another, were localized exclusively in the hypothalamus and showed a distribution pattern similar to that of TH-IR ones. They were distributed in the posterior, dorsal and periventricular hypothalamic areas. Arcuate cells showed no or very weak DA-immunoreactivity. Weak to intense DOPA-IR cells were distributed in a similar manner to DA-IR ones but were far smaller in number. In colchicine-treated animals, DA- and DOPA-immunoreactivities were enhanced particularly in arcuate and dorsal hypothalamic cells. A cluster composed of small DA- and DOPA-IR cells was identified in the area ventral to the mamillothalamic tract equivalent to rat A13c TH-IR cell group. Colchicine treatment enabled us to visualize a large number of TH-IR perikarya in the medial and lateral preoptic areas, anterior commissure nucleus, basal forebrain, area closely related to the organum vasculosum laminae terminalis, and some in the bed nucleus of the stria terminalis as has been reported in other species. However, virtually none of these cells contained detectable DA- and DOPA-immunoreactivities.

DISTRIBUTION OF PROLACTIN RECEPTORS IN THE RAT FOREBRAIN : IMMUNOHISTOCHEMICAL STUDY

The distribution of prolactin receptors (PRL-R) in the rat brain was investigated for the first time with the immunohistochemical technique using monoclonal antibodies raised against PRL-R purified from rat liver. Granular immunostaining was observed in neurons and along their dendritic processes and fibers. PRL-R like immunoreactive neurons were found in a number of brain areas. There was a very dense labelling in the cerebral cortex (pyramidal cell layer), septal nuclei, amygdaloid complex as well as in the hypothalamus (suprachiasmatic, supraoptic, paraventricular and dorsomedial nuclei). A dense staining was seen in the substantia nigra, habenula and in the paraventricular thalamic nucleus. Immunostaining was also found in the choroid plexus and in the subcommissural organ. Comparison between the present distribution and that of PRL-like immunoreactivity indicates that the density of PRL-R generally corresponds to that of the fibers. However, in some regions densely stained by PRL-R antibody, there are very few PRL-immunoreactive fibers. These results are suggestive of different modes of action of PRL in the brain.

Localization of candidate genomic regions influencing paradoxical sleep in mice

QUANTITATIVE trait loci (QTL) approach was used in CXB recombinant inbred mice for preliminary identification of candidate regions on the mouse genome that influence sleep. The only provisional QTLs identified were associated with paradoxical sleep (PS). PS during the light period was associated with markers on chromosome 7 between 7 and 20 centimorgan from the centromere. For PS during the dark period, a single QTL was identified on chromosome 5, near the Clock gene. The 24 h amount of PS was influenced by markers on chromosomes 2, 17, and 19. This first QTL mapping study strongly suggests that a complex behaviour like PS can be controlled by only a few genes.

Demonstration of l-dopa decarboxylating neurons specific to human striatum

In the human striatum, we immunohistochemically demonstrated many neurons that were immunoreactive for aromatic L-amino acid decarboxylase (AADC; the second step dopamine-synthesizing enzyme) but not for tyrosine hydroxylase (TH; the first step dopamine-synthesizing enzyme). The number of AADC-positive neurons was especially large in the ventral striatum including the nucleus accumbens. The significance of AADC-positive neurons are discussed in relation to the acting sites of L-dopa and antipsychotic drugs.

Comparative topography of dopamine- and tyrosine hydroxylase-immunoreactive neurons in the rat arcuate nucleus

The distribution of dopamine (DA)-immunoreactive (IR) cells is described in the rat arcuate nucleus of the hypothalamus and its adjacent areas and compared with that of tyrosine hydroxylase (TH)-IR cells. Small DA-IR cells were seen to be aggregated mainly in the dorsomedial part of the nucleus, but were hardly detectable in its ventrolateral portion and neighbouring periarcuate region which showed many larger TH-IR cells. This study reveals, for the first time, the differences in the respective topography of those neurons which actually contain detectable DA and those which contain TH, the initial synthesizing enzyme of catecholamine.

Neurotoxic lesion of the mesencephalic reticular formation and/or the posterior hypothalamus does not alter waking in the cat

In order to re-evaluate the role of two putative waking systems, we injected a neural cell body toxin, ibotenic acid (IA) (45 micrograms/microliters), into the mesencephalic reticular formation (MRF) and/or the posterior hypothalamus (PH). On the one hand, when the cell body destruction was only restricted to the MRF, the IA microinjection was followed by a temporary high voltage and slow neocortical electroencephalogram (EEG) during the first 24 postoperative hours and by a subsequent long term increase in waking which lasted 8-12 h. After the first postoperative day, there were no motor disturbances, no aphagia nor adypsia, no alteration of cortical activation and no modification of thermoregulation or of the sleep-waking cycle. On the other hand, the IA microinjection into the PH induced a hypothermia during the first postoperative night and a dramatic transient hypersomnia immediately after the disappearance of the anesthesia (14-24 h after the IA injection). On the third day, all cats recovered control level of paradoxical sleep (PS), slow wave sleep (SWS) and cerebral temperature. They presented normal motor behavior but they were not able to eat by themselves during the first postoperative week. Finally, when the lesions of the MRF and the PH were realized in one single operation, the cats were first motionless in a comatose state for 2-3 days. This state was accompanied by a transitory hypothermia and the suppression of a spontaneous or evoked cortical low voltage fast activity. However, from the 2nd postoperative week, both behavioral and EEG waking re-occurred. By contrast, the two successive operations (MRF followed by PH) did not induce a comatose state. We did not observe any deficit in motor behavior, and the sleep-waking cycle was quite normal as from the second postlesion day. In the MRF-PH-lesioned cats, the injection of alpha-methyl-p-tyrosine (150 mg/kg) induced a large decrease in waking and a moderate increase in PS. In the MRF-lesioned cats, IA produced a large area of cell body loss, centered in the MRF, that extended from levels A2 to A6 of stereotaxic planes and sometimes encroached upon the red nucleus and the substantia nigra. In the PH-lesioned cats, the histological analysis revealed a great loss of cell bodies in the PH extended from levels A8 to A12.5. The damage included the lateral and posterior hypothalamic areas and sometimes the tuberomamillary nucleus. In MRF- and PH-lesioned cats, the cell body loss extended from levels A2 to A12.5.(ABSTRACT TRUNCATED AT 400 WORDS)

Origin of the dopaminergic innervation of the rat dorsal raphe nucleus

THE aim of the present study was to describe the distribution of dopamine (DA) fibres in the dorsal raphe nucleus (DRN) and to determine their neurones of origin. Using an anti-DA antibody, we observed a moderate density of DA varicose fibres over the DRN and a dense plexus of DA fibres in the ventrolateral central grey. With a sensitive retrograde tracing technique combining the use of cholera toxin subunit b with tyrosine hydroxylase immunohistochemistry, after tracer injections in the DRN, a few double-labelled cells were observed in the ventral tegmental area and the A10 dorsocaudal DA cell group, as already described. In addition, a moderate number of double-labelled cells was seen in the All hypothalamic DA cell group.

Effect of ambient temperature on the sleep-waking cycle in two strains of mice

The sleep-waking cycle was studied in two strains of mice during continuous exposure over 3 weeks to an ambient temperature of 34 degrees C or 10 degrees C. As compared to the 25 degrees C control period, C57BL/6 mice showed at 34 degrees C a mean increase of 25% in slow-wave sleep (SWS) particularly during the night. Paradoxical sleep (PS) was also increased by 58% during the exposure period particularly during the night and this resulted from an increase in the number of PS episodes. At 10 degrees C SWS was decreased by 9% only during the first week of exposure, whereas PS was reduced by 24% throughout due to a decrease in the number of PS episodes, although their duration increased. The acrophase of the circadian rhythms was not affected by the changes in ambient temperature. In the BALB/c strain at 34 degrees C SWS and PS were increased only during the first week by 13% and 77%, respectively. At 10 degrees C SWS was decreased by 7% throughout and PS was decreased by 36% due to a reduction in the number of PS phases during the light period. On returning to 25 degrees C, a rebound in PS confirmed the PS deprivation in the cold in both strains.