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Effect of behavioral activity on hemodynamic responses induced by whisker stimulation in awake mice
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Abstracts / Neuroscience Research 68S (2010) e109–e222
dition and the CPT, it was shown that to keep the skin in good condition gives a possible effectiveness for the prevention of decline in cutaneous sensation. doi:10.1016/j.neures.2010.07.2270
P1-i11 Nitric oxide is one of major mediators of the relationship between hypoxia and cerebral blood flow increase in rats
Tetsuya Matsuura 1,2 , Hiroyuki Takuwa 2 , Rumiana Bakalova 2 , Takayuki Obata 2 , Iwao Kanno 2 1
Acad Group of Math and Natural Sci, Iwate Univ, Morioka, Japan 2 Molecular Imaging Center, Nat Inst of Radiol Sci, Chiba, Japan
It is known that the regulation of baseline cerebral blood flow (CBF) is correlates with the metabolic demand of oxygen in brain tissue. However, the relationship between the increase in local CBF, induced by neuronal activation (evoked CBF), and oxygen demand in the activated brain area is still a disputable and controversial issue. In the present study, we investigated the effect of hypoxia on the baseline and evoked CBF using laser-Doppler flowmetry (LDF) to clarify the regulation mechanism of CBF in relation to the metabolic oxygen demand in rats. The effect of nitric oxide (NO) synthase inhibition on CBF regulation under hypoxia was also demonstrated using Nω -nitro-L-arginine (LNA). The CBF response to hind-paw stimulation was measured by laser-Doppler flowmetry. Physiological variables, such as heart rate, mean arterial blood pressure (MABP) and PaCO2 during hypoxia, were identical to those during normoxia. In the absence of LNA, hypoxia was accompanied by increase of the baseline CBF and the peak-amplitude of normalized evoked CBF relative to that during normoxia. The peak amplitude of normalized evoked CBF was 20.4±7.4% during hypoxia and 15.3±5.4% during normoxia (P < 0.05). The systemic administration of LNA (hypoxia with LNA) induced a significant enhancement of MABP and reduction of the heart rate in comparison with the values of both parameters, determined under normoxia (P < 0.01). The injection of LNA subjected to hypoxia showed the same baseline CBF as during normoxia, and there is a no significant difference in the normalized evoked CBF between hypoxia with LNA and normoxia. Field potential was constant for all experimental conditions. These results suggest that NO is involved in the enhancement of baseline and evoked CBF during hypoxia. NO inhibitor abolished the effect of hypoxia completely at the baseline level and evoked CBF; therefore, NO could be considered a major mediator of the relationship between hypoxia and evoked CBF response. doi:10.1016/j.neures.2010.07.2272
P1-i12 Morphological analysis of mechanoreceptors in the rat skin identified by intraganglionic injections of neuronal tracers Satomi Ebara 1 , Knarik Bagdasarian 2 , Takahiro Fruta 3 , Taro Koike 4 , Inbal Meir 2 , Ehud Ahissar 2 , Kenzo Kumamoto 1 1
Dept of Anatomy, Meiji Univ of Integrative Medicine, Kyoto 2 Dept of Neurobiology, The Weizmann Institute of Science, Rehovot, Israel 3 Dept of Morphological Brain Science, Kyoto Univ, Kyoto 4 Dept of Anatomy and Cell Science, Kansai Medical Univ, Osaka The aim of this morphological study is to elucidate how single primary sensory neuron innervates skin mechanoreceptors. We investigated the axons labeling after injections of neuronal tracers in trigeminal ganglia (TG) and L5-dorsal root ganglia (DRG) of the rat. TG or DRG were exposed unilaterally in deeply anesthetized adult rats. The TG was visually observed through an opening in a temporal bone of the skull and suction of brain tissues obliquely in the medioventral direction. The dura on the surface of the ganglion was exposed, neuronal tracer (e.g., neurobiotin (NB); 5-20% in 0.1M KCl) was injected in the different parts of the ganglion by a glass pipette or a Hamilton microsyringe. Animals were fixed after 2-20 h survival time after injection. NB-positive neurons were visualized with fluorescence conjugated streptavidin. Relevant tissue was processed using nerve-fiber-teasing method and immunohistochemistry with primary antibodies to pan-axonal marker, PGP9.5 and to myelin basic protein. Labeled single axons were individually and three-dimensionally traced continuously up to the mechanoreceptors in the mystacial pad or foot skin by using confocal microscopic analysis. Five hours of survival time after injection of NB to the TG, was enough to recognize axons in the maxillary nerve at the distance of 20mm. Seven days after injection of PHA-L in TG without suction of brain tissues, the single axon innervating 2 groups of Merkel endings was revealed at the ring sinus of a vibrissal follicle. Another single, not branching axon originated from
L5-DRG was thoroughly traced up to 60mm until to the 5th toe hairy skin. The axon innervated only three neighboring guard hair follicles as palisade endings that were composed of longitudinally arranged lanceolate endings. This study morphologically demonstrated that a primary sensory afferent innervates only one kind of mechanoreceptors at the restricted area. doi:10.1016/j.neures.2010.07.2273
P1-i13 Biphasic plasticity induced by whisker trimming in thalamocortical slices of the mouse barrel cortex Kenji Watanabe Shibuki
, Daiki
Kamatani, Ryuichi
Hishida, Katsuei
Niigata University Brain Research Institute Department of Neurophysiology Endogenous green fluorescence derived from mitochondrial flavoproteins is applicable for functional brain imaging. Using this technique, we investigated cortical responses elicited by thalamic stimulation in the thalamocortical slices including the barrel cortex of mice. Neural activities were observed using patch clamp recording in the areas with a fluorescence response, while almost no activity was found in the areas with no fluorescence response. We further investigated experience-dependent plasticity after whisker trimming. The fluorescence responses were variable from slice to slice. By changing the position of ROIs and stimulating sites systemically, we succeeded to reduce the variability of cortical responses. Whisker trimming during 4-8 weeks or 6-8 weeks of age significantly reduced the responses in the barrel cortex obtained from the mice at 8 weeks old. We thought that whisker trimming with a longer period might more markedly induce this depression. However, there was no clear depression after whisker trimming during 0-8 weeks or 2-8 weeks. These discrepant effects of whisker trimming may be explained by biphasic plasticity induced by whisker trimming: earlier depression caused by loss of activities and later hypersensitivity that compensates the loss of activities. To test this hypothesis, we investigated the effects of transient whisker trimming during 0-7 or 0-6 weeks followed by regrowth of whiskers for 1-2 weeks. Clear potentiation of cortical responses was observed in the mice with transient whisker trimming compared with those of naive mice, supporting the presence of biphasic plasticity in the mouse barrel cortex. doi:10.1016/j.neures.2010.07.2274
P1-i14 Effect of behavioral activity on hemodynamic responses induced by whisker stimulation in awake mice Hiroyuki Takuwa 1 , Joonas Autio 1,2 , Kazuto Takayuki Obata 1 , Iwao Kanno 1 1
Masamoto 1,3 ,
NIRS, Chiba, Japan 2 University of Kuopio, Kuopio, Finland Electro-Communications, Tokyo, Japan
3
University of
In this study, we measured cerebral blood flow (CBF) in awake mice with laser-Doppler flowmetry (LDF). A total of fifteen male C57BL/6J mice were prepared for attaching a custom-made metal head plate to the skull with dental acrylic under isoflurane anesthesia. One day after the surgery, the mice were tethered by screwing the head plate onto a metal rod. Below the animals, a styrofoam ball supported by a jet of air was set in the apparatus. The styrofoam ball freely rotated while the mice walked on it. Regional CBF in the somatosensory cortex and an amount of walking were concurrently monitored with L DF and an optical motion sensor that detect a rotation distance of the ball, respectively. Whisker stimulation (frequency 10 Hz and duration 20 s) was induced a contra-lateral side of the measurement site. Firstly, whisker stimulation-induced blood flow response and behavior were measured during daytime and nighttime. The blood flow responses showed no significant differences between daytime and nighttime (24% and 23% of the pre-stimulus baseline, respectively), whereas the amount of walking was 22% higher under nighttime relative to daytime. Secondly, the blood flow response and behavior were measured from the same single animal over 7days. The mean coefficient of variance among multiple days was observed to be 0.11 (0.08–0.16) and 0.84 (0.52 t o 1.51) for blood flow response and amount of walking, respectively. These results indicate that blood flow response is a relatively stable and reproducible between daytime and nighttime and across multiple days, although amount of walking shows large variations. In conclusion, our system is feasible for a longitudinal study in awake animals. doi:10.1016/j.neures.2010.07.2275
Abstracts / Neuroscience Research 68S (2010) e109–e222
dition and the CPT, it was shown that to keep the skin in good condition gives a possible effectiveness for the prevention of decline in cutaneous sensation. doi:10.1016/j.neures.2010.07.2270
P1-i11 Nitric oxide is one of major mediators of the relationship between hypoxia and cerebral blood flow increase in rats
Tetsuya Matsuura 1,2 , Hiroyuki Takuwa 2 , Rumiana Bakalova 2 , Takayuki Obata 2 , Iwao Kanno 2 1
Acad Group of Math and Natural Sci, Iwate Univ, Morioka, Japan 2 Molecular Imaging Center, Nat Inst of Radiol Sci, Chiba, Japan
It is known that the regulation of baseline cerebral blood flow (CBF) is correlates with the metabolic demand of oxygen in brain tissue. However, the relationship between the increase in local CBF, induced by neuronal activation (evoked CBF), and oxygen demand in the activated brain area is still a disputable and controversial issue. In the present study, we investigated the effect of hypoxia on the baseline and evoked CBF using laser-Doppler flowmetry (LDF) to clarify the regulation mechanism of CBF in relation to the metabolic oxygen demand in rats. The effect of nitric oxide (NO) synthase inhibition on CBF regulation under hypoxia was also demonstrated using Nω -nitro-L-arginine (LNA). The CBF response to hind-paw stimulation was measured by laser-Doppler flowmetry. Physiological variables, such as heart rate, mean arterial blood pressure (MABP) and PaCO2 during hypoxia, were identical to those during normoxia. In the absence of LNA, hypoxia was accompanied by increase of the baseline CBF and the peak-amplitude of normalized evoked CBF relative to that during normoxia. The peak amplitude of normalized evoked CBF was 20.4±7.4% during hypoxia and 15.3±5.4% during normoxia (P < 0.05). The systemic administration of LNA (hypoxia with LNA) induced a significant enhancement of MABP and reduction of the heart rate in comparison with the values of both parameters, determined under normoxia (P < 0.01). The injection of LNA subjected to hypoxia showed the same baseline CBF as during normoxia, and there is a no significant difference in the normalized evoked CBF between hypoxia with LNA and normoxia. Field potential was constant for all experimental conditions. These results suggest that NO is involved in the enhancement of baseline and evoked CBF during hypoxia. NO inhibitor abolished the effect of hypoxia completely at the baseline level and evoked CBF; therefore, NO could be considered a major mediator of the relationship between hypoxia and evoked CBF response. doi:10.1016/j.neures.2010.07.2272
P1-i12 Morphological analysis of mechanoreceptors in the rat skin identified by intraganglionic injections of neuronal tracers Satomi Ebara 1 , Knarik Bagdasarian 2 , Takahiro Fruta 3 , Taro Koike 4 , Inbal Meir 2 , Ehud Ahissar 2 , Kenzo Kumamoto 1 1
Dept of Anatomy, Meiji Univ of Integrative Medicine, Kyoto 2 Dept of Neurobiology, The Weizmann Institute of Science, Rehovot, Israel 3 Dept of Morphological Brain Science, Kyoto Univ, Kyoto 4 Dept of Anatomy and Cell Science, Kansai Medical Univ, Osaka The aim of this morphological study is to elucidate how single primary sensory neuron innervates skin mechanoreceptors. We investigated the axons labeling after injections of neuronal tracers in trigeminal ganglia (TG) and L5-dorsal root ganglia (DRG) of the rat. TG or DRG were exposed unilaterally in deeply anesthetized adult rats. The TG was visually observed through an opening in a temporal bone of the skull and suction of brain tissues obliquely in the medioventral direction. The dura on the surface of the ganglion was exposed, neuronal tracer (e.g., neurobiotin (NB); 5-20% in 0.1M KCl) was injected in the different parts of the ganglion by a glass pipette or a Hamilton microsyringe. Animals were fixed after 2-20 h survival time after injection. NB-positive neurons were visualized with fluorescence conjugated streptavidin. Relevant tissue was processed using nerve-fiber-teasing method and immunohistochemistry with primary antibodies to pan-axonal marker, PGP9.5 and to myelin basic protein. Labeled single axons were individually and three-dimensionally traced continuously up to the mechanoreceptors in the mystacial pad or foot skin by using confocal microscopic analysis. Five hours of survival time after injection of NB to the TG, was enough to recognize axons in the maxillary nerve at the distance of 20mm. Seven days after injection of PHA-L in TG without suction of brain tissues, the single axon innervating 2 groups of Merkel endings was revealed at the ring sinus of a vibrissal follicle. Another single, not branching axon originated from
L5-DRG was thoroughly traced up to 60mm until to the 5th toe hairy skin. The axon innervated only three neighboring guard hair follicles as palisade endings that were composed of longitudinally arranged lanceolate endings. This study morphologically demonstrated that a primary sensory afferent innervates only one kind of mechanoreceptors at the restricted area. doi:10.1016/j.neures.2010.07.2273
P1-i13 Biphasic plasticity induced by whisker trimming in thalamocortical slices of the mouse barrel cortex Kenji Watanabe Shibuki
, Daiki
Kamatani, Ryuichi
Hishida, Katsuei
Niigata University Brain Research Institute Department of Neurophysiology Endogenous green fluorescence derived from mitochondrial flavoproteins is applicable for functional brain imaging. Using this technique, we investigated cortical responses elicited by thalamic stimulation in the thalamocortical slices including the barrel cortex of mice. Neural activities were observed using patch clamp recording in the areas with a fluorescence response, while almost no activity was found in the areas with no fluorescence response. We further investigated experience-dependent plasticity after whisker trimming. The fluorescence responses were variable from slice to slice. By changing the position of ROIs and stimulating sites systemically, we succeeded to reduce the variability of cortical responses. Whisker trimming during 4-8 weeks or 6-8 weeks of age significantly reduced the responses in the barrel cortex obtained from the mice at 8 weeks old. We thought that whisker trimming with a longer period might more markedly induce this depression. However, there was no clear depression after whisker trimming during 0-8 weeks or 2-8 weeks. These discrepant effects of whisker trimming may be explained by biphasic plasticity induced by whisker trimming: earlier depression caused by loss of activities and later hypersensitivity that compensates the loss of activities. To test this hypothesis, we investigated the effects of transient whisker trimming during 0-7 or 0-6 weeks followed by regrowth of whiskers for 1-2 weeks. Clear potentiation of cortical responses was observed in the mice with transient whisker trimming compared with those of naive mice, supporting the presence of biphasic plasticity in the mouse barrel cortex. doi:10.1016/j.neures.2010.07.2274
P1-i14 Effect of behavioral activity on hemodynamic responses induced by whisker stimulation in awake mice Hiroyuki Takuwa 1 , Joonas Autio 1,2 , Kazuto Takayuki Obata 1 , Iwao Kanno 1 1
Masamoto 1,3 ,
NIRS, Chiba, Japan 2 University of Kuopio, Kuopio, Finland Electro-Communications, Tokyo, Japan
3
University of
In this study, we measured cerebral blood flow (CBF) in awake mice with laser-Doppler flowmetry (LDF). A total of fifteen male C57BL/6J mice were prepared for attaching a custom-made metal head plate to the skull with dental acrylic under isoflurane anesthesia. One day after the surgery, the mice were tethered by screwing the head plate onto a metal rod. Below the animals, a styrofoam ball supported by a jet of air was set in the apparatus. The styrofoam ball freely rotated while the mice walked on it. Regional CBF in the somatosensory cortex and an amount of walking were concurrently monitored with L DF and an optical motion sensor that detect a rotation distance of the ball, respectively. Whisker stimulation (frequency 10 Hz and duration 20 s) was induced a contra-lateral side of the measurement site. Firstly, whisker stimulation-induced blood flow response and behavior were measured during daytime and nighttime. The blood flow responses showed no significant differences between daytime and nighttime (24% and 23% of the pre-stimulus baseline, respectively), whereas the amount of walking was 22% higher under nighttime relative to daytime. Secondly, the blood flow response and behavior were measured from the same single animal over 7days. The mean coefficient of variance among multiple days was observed to be 0.11 (0.08–0.16) and 0.84 (0.52 t o 1.51) for blood flow response and amount of walking, respectively. These results indicate that blood flow response is a relatively stable and reproducible between daytime and nighttime and across multiple days, although amount of walking shows large variations. In conclusion, our system is feasible for a longitudinal study in awake animals. doi:10.1016/j.neures.2010.07.2275