IMMUNOHISTOCHEMICAL IDENTIFICATION AND DISTRIBUTION OF GLUTAMATERGIC NMDA AND mGlu1 RECEPTORS IN THE PONTINE INTERTRIGEMINAL REGION IN RATS

M. Stoiljkovic
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Abstract

Local glutamate simulation of intertrigeminal region (ITR) in the lateral pons evoked immediate cardiovascular and respiratory effects proposing its role in central cardiorespiratory control. Since pharmacological studies provided only functional evidence for the existence of glutamate receptors in the ITR and thereby specifying putative neurochemical substrate involved in this control, here we employed immunohistochemistry to examine expression and distribution of NMDA and mGlu1 receptors in this structure. Thirty adult male Sprague-Dawley rats were perfuse-fixed, their brains frozen and cut into sequential series of 20 µm thick sections through the ITR. Immunohistochemistry was performed using polyclonal antibodies against NMDA-NR1, NMDA-NR2A and mGlu1 receptors. Labeled neurons in the ITR were analyzed using light microscope and computerized image analysis system for quantification of relative immunoreactivity as the mean of integrated optical density (IOD), and counting the immunopositive cells. Light microscopic analyses demonstrated NMDA-NR1-immunoreactivity mainly localized in the neuronal cell bodies with sparse distribution on primary dendrites, while NMDA-NR2A-immunoreactivity was basically somatically distributed. The mGlu1-immunoreactivity was moderate and observed both in neuronal bodies and primary dendrites or extracellular matrix suggesting somatodendritic localization. Quantitative analyses of IOD showed very strong expression of NMDA-NR1, weak of NMDA-NR2A and strong-to-moderate expression of mGluR1, with differences in immunostaining signal distribution over rostro-caudal span of the ITR. Counting of immunopositive cells followed similar expression profile. Our data directly confirm the presence of glutamatergic NMDA and mGlu1 receptors in the ITR apparently involved in signaling pathways by which this region modulates cardiorespiratory functions such as blood pressure, heart rate and breathing.
大鼠脑桥三叉间区谷氨酸能NMDA和mGlu1受体的免疫组织化学鉴定和分布
脑桥外侧三叉间区(ITR)的局部谷氨酸模拟引起了立即的心血管和呼吸作用,表明其在中枢心肺控制中的作用。由于药理学研究仅提供了ITR中谷氨酸受体存在的功能证据,从而确定了参与该控制的推定神经化学底物,因此我们采用免疫组织化学方法检测了NMDA和mGlu1受体在该结构中的表达和分布。将30只成年雄性Sprague-Dawley大鼠灌注固定,将其大脑冷冻,并通过ITR切成20µm厚的连续系列切片。采用抗NMDA-NR1、NMDA-NR2A和mGlu1受体的多克隆抗体进行免疫组化。利用光镜和计算机图像分析系统对ITR中标记的神经元进行定量分析,以相对免疫反应性作为积分光密度(IOD)的平均值,对免疫阳性细胞进行计数。光镜分析显示nmda - nr1免疫反应性主要集中于神经元细胞体,在初生树突上分布稀疏,而nmda - nr2a免疫反应性基本呈体分布。mglu1的免疫反应性是中等的,在神经元体和初级树突或细胞外基质中都观察到,提示体树突定位。定量分析IOD显示NMDA-NR1的表达非常强,NMDA-NR2A的表达较弱,mGluR1的表达较强至中等,在ITR的上尾跨上免疫染色信号分布存在差异。免疫阳性细胞计数遵循类似的表达谱。我们的数据直接证实了谷氨酸能NMDA和mGlu1受体在ITR中的存在,这些受体显然参与了该区域调节血压、心率和呼吸等心肺功能的信号通路。
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