Neuroglial Reaction in the Substantia Nigra and Striatum of 6-Hydroxydopamine Induced Parkinson's Disease Rat Model

OBJECTIVES: Parkinson's disease is a well-known neurodegenerative disease characterized by dopaminergic cell death in the substantia nigra. The reactive gliosis by activated astrocytes and microglias is no more regarded as a simple sequel of neuronal cell death. Microglial activation takes place in a stereotypic pattern with graded morphologic and functional(resting, activated and phagocytic) changes. In Parkinson's disease animal model, the degree of microglial activation along the nigro-striatal dopaminergic tract has not been studied intensively. The purpose of this study was to elucidate the characteristics of microglial reaction and to grade its degree of activation at substantia nigra and corpus striatum using 6-hydroxydopamine induced rat model of Parkinson's disease. METHODS: Using Sprague-Dawley rat, parkinsonian model was made by 6-hydroxydopamine(OHDA) induced destruction of medial and lateral substantia nigra(SN). The rat was sacrificed 3-, 5-, 7-, 14- and 21-day-after operation. For control group, we injected saline with same manner and sacrificed 3-day after operation. With immunohistochemistry, we examined dopaminergic neuronal cells and microglial expression using tyrosine hydroxylase (TH) and OX-42 antibodies, respectively. Also we performed in situ hybridization for osteopontin, a possible marker of subset in activated microglia. RESULTS: 1) In lesioned side of substantia nigra and corpus striatum, the TH immunoreactivity was markedly decreased in whole experimental groups. 2) Using optical densitometry, microglia induced immunoreactivity of OX-42 was counted at SN and corpus striatum. At SN, it was increased significantly on the lesioned side in control and all time-dependent experimental groups. At striatum, it was increased significantly in post lesion 3-day group only(p < 0.05). Compared to control group, immunoreactivity of OX-42 on lesioned side was increased in groups, except post lesion 21-day group, at SN. Only post lesion 3-day group showed significance at striatum(p < 0.05). Compared to SN region, immunoreactivity of OX-42 was much weaker in striatum. 3) Microscopically, the microglias showed typically different activation pattern. At SN, numerous phagocytic microglias were found at pars compacta and reticularis of lesion side. At striatum, no phagocytic form was found and the intensity of staining was much weaker. 4) At SN, the immunoreactivity of osteopontin showed definite laterality and it was markedly increased at pars compacta of lesion side with relatively short duration time. At striatum, however, it was not detected by in situ hybridization technique. CONCLUSION: The nigral 6-OHDA induced rat model of Parkinson's disease revealed several characteristic patterns of microglial reaction. At SN, microglias was activated shortly after direct neuronal damage and maintained for about three weeks. In contrast, despite of sufficient dopaminergic insufficiency at striatum, activation of microglias was trivial, and distinguished 3 day later. Antegrade slow neuronal degeneration is major pathophysiology in striatal dopaminergic deficiency. So, the acuteness of neuronal damage and consequential degree of neuronal degeneration may be important factor for microglial activation in neurodegenerative diseases such as Parkinson's disease. Additionally, osteopontin may be a possible marker for several subsets of activated microglia, possibly the phagocytic form.

Alterations of Heart Rate Variability by Vestibular Stimulation in Rabbits

BACKGROUND: There is a substantial evidence that anatomical connections and functional interactions exist between vestibular and autonomic systems. The nature of these interactions, however, is complex and has not been fully defined. Heart rate variability (HRV) was used to investigate the physiological role of the vestibular system on control of heart rate. METHODS: HRV including mean, standard deviation, coefficient of variation (CV), power spectrum was analyzed from R-R intervals of ECG during vestibular stimulation in rabbits. RESULTS: Urethane anesthesia increased heart rate and maintained regular R-R intervals, however, low frequency region/high frequency region (LF/HF) was not changed. In anesthetized rabbits, electrical stimulation of the vagus nerve decreased heart rate and decreased LF/HF by increasing HF. On the contrary, electrical stimulation of the cervical sympathetic nerve increased heart rate and increased LF/HF by increasing LF. Atropine, cholinergic blocker, increased heart rate and increased LF/HF by reducing HF, and propranolol, beta-adrenergic blocker, decreased heart rate and decreased LF/HF by reducing LF. In unanesthetized rabbits, stimulation of the vestibular system induced by rotation or caloric increased heart rate and increased LF/HF by increasing LF. Also electrical stimulation of the vestibular nerve produced the same effects as rotation or caloric in anesthetized rabbits. CONCLUSION: These results suggest that stimulation of the vestibular system increased heart rate not by inhibiting the parasympathetic nerve but by activating the sympathetic nerve.

Role of Vestibulosympathetic Reflex on Orthostatic Hypotension in Rats

BACKGROUND: The orthostatic hypotension in response to the assumption of an upright posture is regulated by activation of sympathetic nerves. Role of the vestibular system and neural pathway on orthostatic hypotension were investigated. METHODS: Changes of arterial blood pressure produced by head-up tilting, rotatory stimulation of the vestibular system, or electrical stimulation to the vestibular nerve, vestibular nuclei, and rostral ventrolateral medulla (RVLM) were measured in Sprague-Dawley rats. Also, field potentials were recorded in the vestibular nuclei and RVLM and c-Fos expression was evaluated in the brain stem in order to investigate the vestibulosympathetic pathways. RESULTS: The three phasic blood pressure responses were elicited by head-up tilting: initial fall, early recovery, and late sustained pressure at near control levels, the magnitude of the pressure fall was parallel with the degree of head-up tilting in normal rats. Return position from head-up tilting recovered control level of blood pressure after a brief rapid elevation. However, bilateral labyrinthectomy resulted in exaggerated initial falling and devoid of early recovery phase during postural change. Sinusoidal rotation about off-vertical axis of the vestibular system elicited more elevation of blood pressure than rotation about earth vertical axis. Electrical stimulation of the vestibular nerve, vestibular nucleus, and RVLM produced elevation of blood pressure, which was the most prominent by stimulation of RVLM. Field potentials composed of P, N1, N2 waves in the vestibular nuclei were recorded by stimulation of the vestibular nerve, while weak potentials in RVLM were recorded by stimulation of the vestibular nuclei. An electrical stimulation of the vestibular nuclei expressed c-Fos immunoreactive cells in RVLM. CONCLUSION: These results suggest that the otolith organ of the vestibular system plays a major role in control of orthostatic hypotension, and the pathway of vestibulosympathetic reflex in control of blood pressure involves the vestibular nuclei, RVLM, intermed-iolateral nuclei of the thoracic spinal cord.

Effects of electrical stimulation of the vestibular system on neuronal activity of the ipsilateral medial vestibular nuclei following unilateral labyrinthectomy in rats

The purpose of this study was to evaluate the effects of electrical stimulation on vestibular compensation following ULX in rats. Electrical stimulation (ES) with square pulse (100 ~ 300 uA, 1.0 ms, 100 Hz) was applied to ampullary portion bilaterally for 6 and 24 hours in rats receiving ULX. After ES, animals that showed the recovery of vestibular symptoms by counting and comparing the number of spontaneous nystagmus were selected for recording resting activity of type I, II neurons in the medial vestibular nuclei (MVN) of the lesioned side. And then the dynamic neuronal activities were recorded during sinusoidal rotation at a frequency of 0.1 Hz and 0.2 Hz. The number of spontaneous nystagmus was significantly different 24 hours (p< 0.01, n = 10), but not 6 hours after ULX+ES. As reported by others, the great reduction of resting activity only in the type I neurons ipsilateral to lesioned side was observed 6, 24 hours after ULX compared to that of intact labyrinthine animal. However, the significant elevation (p < 0.01) of type I and reduction (p < 0.01) of type II neuronal activity were seen 24 hours after ULX+ES. Interestingly, gain, expressed as maximum neuronal activity(spikes/sec)/maximum rotational velocity (deg/sec), was increased in type I cells and decreased in type II cells 24 hours after ULX+ES in response to sinusoidal rotation at frequencies of both 0.1 Hz and 0.2 Hz. This result suggests that accompanying the behavioral recovery, the electrical stimulation after ULX has beneficial effects on vestibular compensation, especially static symptoms (spontaneous nystagmus), by enhancing resting activity of type I neurons and reducing that of type II neurons.

Effect of electrical stimulation of the vestibular system on vestibuloocular reflex and c-Fos expression in the medial vestibular nuclei of unilateral labyrinthectomized rats

Unilateral labyrinthectomy (ULX) causes autonomic symptoms, ocular and postural asymmetries, which disappear over time in the process of equilibrium recovery known as vestibular compensation. In the present study, in order to elucidate mechanisms responsible for the effects of electrical stimulation on vestibular compensation and investigate the relationship between vestibular compensation and c-Fos expression in the medial vestibular nuclei following ULX, we measured spontaneous nystagmus, eye movement induced by sinusoidal rotation and c-Fos expression up to 72 hs after ULX in Sprague-Dawley rats. Experimental animals were divided into two groups: ULX group with ULX only, and electrical stimulation (ES) group with electrical stimulation of -2 ~ -5 V, 1.0 ms, 100 Hz to the lesioned vestibular system for 4 hs/day. Spontaneous nystagmus following ULX disappeared by 72 hs in ULX group and 36 hs in ES group. In eye movement induced by sinusoidal rotation, normal pattern of eye movement by rotation toward the lesioned side was recovered 24 hs after ULX at rotation of 0.1 Hz and 6 hs after at 0.2 Hz, 0.5 Hz in ULX group. In ES group, the eye movement recovered after 12 hs at 0.1 Hz, 6 hs at 0.2 Hz, and 4 hs at 0.5 Hz. Directional preponderance which represents the symmetry of bilateral vestibular functions showed significantly early recovery in ES group compared with that of ULX group. Expression of c-Fos immunoreactive cells in the bilateral medial vestibular nuclei was severely asymmetrical till 36 hs in ULX group, and then it became a symmetry and disappeared after 72 hs. However, ES group showed the symmetry of c-Fos expression after 6 hs, which was significantly early recovery in ES group. All these findings suggest that electrical stimulation ameliorates recovery of ventibuloocular reflex following ULX by the restoration of the balance of the resting activity between bilateral medial vestibular nuclei. In addition, c-Fos expression in the medial vestibular nuclei could be used as a marker of vestibular compensation since c-Fos expression is closely related to the course of recovery following ULX.

Cell Death and Cell Proliferation during Histogenesis in the Rat Retina after Birth / 대한해부학회지

During development of central nervous system, cell proliferation, cell migration, cell differentiation and cell death are required. It has been reported that a number of cells are dying during development in the mammalian retinae examined so far, but the pattern of cell death has not been clarified yet. In addition. little has been studied on cell proliferation after birth. This study was conducted to identify histogenesis, cell death and cell proliferation in the retinae of the developing rats by light and electron microscopic methods as well as by immunohistochemical method using anti-proliferating cell nuclear antigen [PCNA] antiserum. The results were as follows : 1. In the developing rat, from postnatal 0 through 7 days, retina consisted of ganglion cell layer, inner plexiform layer and neuroblast layer. Neuroblast layer could be subdivided into three sublaminae : sublamina a, sublamina b and sublamina p, from postnatal 3 through 7 days. 2. From postnatal 10 days, retina consisted of ganglion cell layer, inner plexiform layer, inner nuclear layer, outer plexiform layer and outer nuclear layer. 3. Cells undergoing degeneration were observed from postnatal 0 to 13 days, and patterns of cell death were apoptosis, cytoplasmic degeneration and autophagic degeneration. 4. PCNA-immunoreactivity was seen in the cells located in sublaminae b and p of the neuroblast layer at postnatal 0 and 1 days. From postnatal 3 days PCNA immunoreactivity decreased. At 7-day-old rat, PCNA-Immunoreactive cells scattered in the distal part of sublamina p of the neuroblast layer.No immunoreactivity was observed from postnatal 10 days. These results demonstrate that retinal cell proliferation ends at postnatal 7 days, and histogenesis of retina is completed at postnatal 10 days, and superfluous cells during retinal development are eliminated by apoptosis, cytoplasmic degeneration and autophagic degeneration.

Immunocytochemical Study on Synaptic Circuitry of Glycinergic Neurons in the Rat Retina / 대한해부학회지

The role of glycine as an inhibitory neurotransmitter is well established, and glycinergic neurons appear to play an important role in the mammalian retinae[Ikeda & Sheardown, 1983 ; Bolz et al., 1985]. Though it has been reported that certain conventional and displaced amacrine cells and a few of bipolar cells are consistently labeled with anti-glycine antiserum in the mammalian retinae so far[W ssle et al., 1986 ; Pourcho & Goebel, 1987 ; Davanger et al., 1991 ; Yoo & Chung, 1992], little has been studied on the synaptic circuitry of glycinergic neurons to clarify mechanism of its action in the visual processing of the mammalian retinae. This study was conducted to localize glycinergic neurons and to define their synaptic circuitry in the rat retina by immunocytochemical method using anti -glycine antiserum. The results were as follows : 1. Glycinergic neurons of the rat retina were conventional and displaced amacrine cells, interstitial cells and bipolar cells. 2. Glycinergic amacrine cells could be subdivided into two types, that is, A II amacrine cells and other amacrine cells, according to their ultrastructures. Glycinergic A II amacrine and other amacrine cell processes comprised postsynaptic dyad at the ribbon synapse of rod bipolar axon terminals in the sublamina b of the inner plexiform layer of the retina. Glycinprgic A II amacrine cell processes made gap junctions with axon terminals of unlabeled invaginating cone bipolar cells in the sublamina b, and made chemical synapses onto axon terminals of unlabeled flat cone bipolar cells and onto dendrites of ganglion cells in the sublamina a of the inner plexiform layer. In the sublamina b of the inner plexiform layer, g1ycinergic amacrine cell processes were postsynaptic to axon terminals of unlabeled invaginating cone bipolar cells, and made chemical output synapses onto axon terminals of unlabeled invaginating cone bipolar and rod bipolar cells and onto the dendrites of ganglion cells. Such cases that pre- and post-synaptic processes of glycinergic amacrine cell processes were non- glycinergic amacrine cell processes were frequently observed throughout the inner plexiform layer. In some cases, glycinergic amacrine cell processes receiving synaptic inputs from other glycinergic amacrine cell process made synaptic outputs onto the non-glycinergic or glycinergic amacrine cell processes. 3. Glycinergic bipolar cells could be subdivided into invaginating and flat cone bipolar cells. Postsynaptic dyads of cone bipolar cells at the ribbon synapses were non-glycinergic amacrine and amacrine cell processes, glycinergic amacrine and amacrine cell processes, glycinergic amacrine and non-glycinergic amacrine cell processes, and dendrite and dendrite of ganglion cells. These results demonstrate that [1] glycinergic A II amacrine cell receiving synaptic input from rod bipolar cells inhibit flat cone bipolar cells and OFF ganglion cells via chemical synapse, and excite ON cone bipolar cells via electrical synapse ; thereby visual information in the darkness can be transmitted to ON ganglion cells via ON cone bipolar cells, and [2] glycine released from glycinergic neurons inhibits directly ON and OFF ganglion cells or indirectly ON and OFF ganglion cells via non-glycinergic amacrine or bipolar cells.