Yoshiki Takeuchi

Early maternal deprivation induces alterations in brain-derived neurotrophic factor expression in the developing rat hippocampus

The effects of maternal deprivation (MD) during early postnatal life on the brain-derived neurotrophic factor (BDNF) level were investigated in the present study. Wistar rats were assigned to either maternal deprivation or mother-reared control (MRC) groups. MD manipulation was achieved by separating rat pups from their mothers for 3h a day during postnatal days (PND) 10-15. At 16, 20, 30, and 60 days of age, the level of BDNF mRNA in the hippocampal formation of each group was determined using real-time PCR analysis. Early postnatal maternal deprivation of rat pups resulted in a significant increase in body weight at 60 days of age. The expression of BDNF mRNA in the hippocampus was significantly decreased at 16 days of age, and increased at 30 and 60 days of age. These data indicate that even a brief period of maternal deprivation during early postnatal life can affect hippocampal BDNF expression.

The effect of the timing of prenatal X-irradiation on Purkinje cell numbers in rat cerebellum.

Exposure of the developing brain to X-irradiation in utero is known to cause various deleterious consequences. We have previously reported the effects of prenatal X-irradiation on the development of the cerebral cortex in rats. We have now extended this study to examine the effects of such X-irradiation on the development of the cerebellum. Wistar rats were exposed to 1.5 Gy X-irradiation either on days 14, 15 or 16 of gestation (E14, E15, E16). Sham-irradiated animals were used as controls. At seven postnatal weeks of age, male rats from each group were deeply anesthetized and killed by intracardiac perfusion with 2.5% glutaraldehyde in 0.1 M phosphate buffer. The unbiased stereological procedure known as the fractionator method was used to estimate the total number of Purkinje cells in the cerebellum of each animal. Body and cerebellar weights from E14 and E15, but not E16 irradiated rats showed significant deficits compared to control animals. Rats irradiated on E16 and control rats had about 285100-304800 Purkinje cells in the cerebellum. There was no significant difference between these values. However, E14 and E15 irradiated animals had about 117500 and 196300 Purkinje cells, respectively. These estimates were significantly different from those observed in both control and E16 irradiated rats. Given that the phase of division of Purkinje cell progenitors is mainly between E14-E15 and the phase of differentiation and migration is between E16-E20, it is concluded that the vulnerable period of the Purkinje cells to X-irradiation closely overlaps the phase of division of progenitors.

Colocalization of taurine and glial fibrillary acidic protein immunoreactivity in mouse hippocampus induced by short-term ethanol exposure.

Morphological changes of the hippocampus were investigated in mice exhibiting signs of intoxication following short-term exposure to 6% ethanol. These alterations were examined by a double immunofluorescent study using antibodies to taurine and anti-glial fibrillary acidic protein (GFAP) antibody. Antibody-labeled taurine was localized mainly in the astrocytes and endothelial cells of control mice. Ethanol administration resulted in a significant increase in the accumulation of taurine and GFAP immunoreactivity (IR) in the stratum lacunosum-moleculare (sl-m) of the hippocampus. Specifically, the cell bodies of taurine-positive astrocytes were hypertrophied, their processes were elongated in the pericapillary region, and some colocalized with GFAP-IR cells. Furthermore, quantitative analysis revealed that the merged area in ethanol-treated mice was twice that (71.6% vs. 35.8%) of control mice. Since taurine is involved in various neuroprotective functions, the present observations suggest that the expression of taurine IR in reactive astrocytes after ethanol exposure might play an important role in neuroprotective processes.

Afferent fibers in the hypoglossal nerve: A horseradish peroxidase study in the cat

The existence of afferent fibers in the cat hypoglossal nerve was studied by transganglionic transport of horseradish peroxidase (HRP). Injections of wheat germ agglutinin-conjugated HRP (WGA-HRP) into the hypoglossal nerve resulted in some retrograde labeling of cell bodies within the superior ganglia of the ipsilateral glossopharyngeal and vagal nerves. A few labeled cell bodies were also present ipsilaterally within the inferior ganglion of the vagal nerve and the spinal ganglion of the C1 segment. Some of the labeled glossopharyngeal and vagal fibers reached the nucleus of the solitary tract by crossing the dorsal portion of the spinal trigeminal tract. Others distributed to the spinal trigeminal nucleus pars interpolaris and to the ventrolateral part of the medial cuneate nucleus by descending through the dorsal portion of the spinal trigeminal tract. In the spinal cord these descending fibers, intermingling with labeled dorsal root fibers, distributed to laminae I, IV-V and VII-VIII of the C1 and C2 segments. Additional HRP experiments revealed that the fibers in laminae VII-VIII originate mainly from dorsal root of the C1 segment.

Regional differences in vulnerability of the cerebellar foliations of rats exposed to neonatal X-irradiation

The purpose of the present study was to elucidate regional differences in the vulnerability of cerebellar foliations of rats exposed to X-irradiation. Effects of X-irradiation on foliations were examined with respect to histological changes in Purkinje cells and Bergmann glial fibers by calbindin-D28k (CB) and glial fibrillary acidic protein (GFAP) immunohistochemistry, respectively. Wistar rats were exposed to X-irradiation (1.5 Gy) on postnatal day (PND) 1. At 3 weeks of age, the cerebellum was examined. The cerebella of rats exposed to X-irradiation showed smaller and abnormal foliations compared with controls. Fewer cerebellar foliations due to fusion with neighboring folia were observed in folia I-III and VIa-VII. Moreover, the extent of such abnormalities was more severe in the latter folia. CB-immunoreactive (IR) Purkinje cells exhibited thin, short, disoriented dendrites that had invaded the granular layer or white matter. On the other hand, GFAP-IR Bergmann glial fibers had not extended their processes into the molecular layer perpendicular to the pial surface, and they appeared thin and disoriented. Accordingly, the above cerebellar abnormalities were more severe in folia I-III, VIa-VII and X than in other regions. In contrast to the histological alterations in these folia, there were no apparent qualitative differences in folia IV-V between X-irradiated and controls. These findings indicate regional difference in the vulnerability of cerebellar folia to X-irradiation. Such differences might be attributed to the cerebellar neurogenetic gradient.

Multiple defects in the formation of rat cortical axonal pathways following prenatal X-ray irradiation

Prenatal X‐ray irradiation is known to result in severe defects of neuronal migration and laminar formation in the cerebral cortex. We examined the formation of cortical afferent and efferent pathways in rats that had been exposed to X‐ray irradiation (1.0u2003Gy) at embryonic day 14 (E14), by birthdating with bromodeoxyuridine (BrdU) and axonal labeling with 1‐1′‐dioctodecyl‐3,3,3′,3′‐ tetramethyl‐indocarbocyanine perchlorate (DiI), in addition to immunohistochemical staining for various axonal markers including neurofilament, and cell adhesion molecules L1 and TAG‐1. The results obtained were as follows. (i) The neuroepithelium formed germinal rosettes and concavities in the cortical anlage from 2u2003days after irradiation. Neurons generated in the neuroepithelium accumulated to form subcortical heterotopia and obstructed pathway formation in the intermediate zone, resulting in an aberrant trajectory of TAG‐1‐immunoreactive cortical efferent axons. (ii) In rats exposed to X‐ray irradiation at E14, cystic cavities were formed in the cortex–striatum boundary region between E15 and E17, probably because of delayed cell death of neurons generated at E14. These cavities transiently interrupted both cortical afferent (L1‐positive) and efferent (TAG‐1‐positive) axons. (iii) X‐ray irradiation at E14 partially destroyed subplate neurons (transient targets of thalamic afferent axons) and disturbed the arrangement of the subplate layer. This resulted in a misrouting of neurofilament‐ and L1‐immunoreactive thalamocortical axons that obliquely traversed the cortical plate to run up to the superficial layer. The present study demonstrates for the first time that X‐ray irradiation during initial cortical development causes multiple defects in the formation of cortical afferent and efferent pathways.

Excessive testosterone treatment and castration induce reactive astrocytes and fos immunoreactivity in suprachiasmatic nucleus of mice

The suprachiasmatic nucleus (SCN) has long been recognized as the central mammalian circadian pacemaker that controls behavioral and physiological processes. The role of the SCN in circadian rhythms has been the subject of a wide range of physiological and behavioral studies, although the influence of homeostasis rhythms (such as fluctuating hormone levels) on the SCN of the hypothalamus is not entirely clear. The present study was undertaken to examine the morphological interactions between astroglial and neuronal elements in the SCN of mice after either a short-term excessive testosterone treatment (ETT) or castration, using glial fibrillary acidic protein (GFAP), and immediate early gene c-fos as well as calbindin-D28k (CB) immunohistochemistry. Both ETT and castration resulted in a significant increase in the accumulation of reactive astrocytes and Fos-imunoreactivity (IR), especially in the dorsomedial (DM) sub-region of the SCN. However, CB-IR neurons in the examined brain regions showed little change. These findings indicate that the DM sub-region of the SCN may be a possible center of hormonal regulation via a hypothalamic neuroendocrine circuit, and that a non-photic stimuli mechanism might play a role in circadian rhythm regulation.

Parabrachial inputs to Fos-immunoreactive neurons in the lateral central nucleus of amygdala activated by hypotension: a light and electron microscopic study in the rat

Morphological features and functional implications of projections of the parabrachial nucleus to the central nucleus of the amygdala were investigated in the rat. The anatomical study was based on injections of the tracers horseradish peroxidase and biotinylated dextran amine. An extremely dense concentration of labeled fibers was found in the lateral and lateral capsular subdivisions of the central nucleus of the amygdala, originating mainly from the external lateral and ventral lateral subnuclei of the parabrachial nucleus. The parabrachial fibers exhibited the morphological characteristic of forming dense pericellular terminal arborizations. The functional implications of this pathway in cardiovascular functions were verified using Fos protein induction in response to hypotension induced by continuous intravenous administration of hydralazine-hydrochloride. In this paradigm, Fos immunoreactivity was found to be confined to the lateral and lateral capsular subdivisions of the central nucleus of the amygdala. Double immunostaining methods were used to visualize, at the electron microscopic level, terminals labeled by biotinylated dextran amine and Fos cell labeling. With this approach, we were able to confirm that Fos-immunoreactive neurons in the central nucleus of the amygdala receive axosomatic terminals from the parabrachial nucleus. The present findings point out that parabrachial inputs to the central nucleus of the amygdala play a relevant role in regulating cardiovascular function.