Lorier Amanda R, Huxtable Adrianne G, Robinson Dean M, Lipski Janusz, Housley Gary D, Funk Gregory D
Department of Physiology, Faculty of Medicine and Dentistry, University of Alberta, Edmonton, Alberta, Canada T6G 2H7.
J Neurosci. 2007 Jan 31;27(5):993-1005. doi: 10.1523/JNEUROSCI.3948-06.2007.
ATP is released during hypoxia from the ventrolateral medulla (VLM) and activates purinergic P2 receptors (P2Rs) at unknown loci to offset the secondary hypoxic depression of breathing. In this study, we used rhythmically active medullary slices from neonatal rat to map, in relation to anatomical and molecular markers of the pre-Bötzinger complex (preBötC) (a proposed site of rhythm generation), the effects of ATP on respiratory rhythm and identify the P2R subtypes responsible for these actions. Unilateral microinjections of ATP in a three-dimensional grid within the VLM revealed a "hotspot" where ATP (0.1 mM) evoked a rapid 2.2 +/- 0.1-fold increase in inspiratory frequency followed by a brief reduction to 0.83 +/- 0.02 of baseline. The hotspot was identified as the preBötC based on histology, overlap of injection sites with NK1R immunolabeling, and potentiation or inhibition of respiratory frequency by SP ([Sar9-Met(O2)11]-substance P) or DAMGO ([D-Ala2,N-MePhe4,Gly-ol5]-enkephalin), respectively. The relative potency of P2R agonists [2MeSADP (2-methylthioadenosine 5'-diphosphate) approximately = 2MeSATP (2-methylthioadenosine 5'-triphosphate) approximately = ATPgammas (adenosine 5'-[gamma-thio]triphosphate tetralithium salt) approximately = ATP >> UTP approximately = alphabeta meATP (alpha,beta-methylene-adenosine 5'-triphosphate)] and attenuation of the ATP response by MRS2179 (2'-deoxy-N6-methyladenosine-3',5'-bisphosphate) (P2Y1 antagonist) indicate that the excitation is mediated by P2Y1Rs. The post-ATP inhibition, which was never observed in response to ATPgammas, is dependent on ATP hydrolysis. These data establish in neonatal rats that respiratory rhythm generating networks in the preBötC are exquisitely sensitive to P2Y1R activation, and suggest a role for P2Y1Rs in respiratory motor control, particularly in the P2R excitation of rhythm that occurs during hypoxia.
在缺氧期间，三磷酸腺苷（ATP）从延髓腹外侧区（VLM）释放，并激活未知位点的嘌呤能P2受体（P2Rs），以抵消呼吸的继发性缺氧性抑制。在本研究中，我们使用新生大鼠有节律活动的延髓切片，相对于前包钦格复合体（preBötC）（一个推测的节律产生位点）的解剖学和分子标记，绘制ATP对呼吸节律的影响，并确定负责这些作用的P2R亚型。在VLM内的三维网格中单侧微量注射ATP，发现一个“热点”，在此处ATP（0.1 mM）引起吸气频率迅速增加2.2±0.1倍，随后短暂降至基线的0.83±0.02。根据组织学、注射部位与神经激肽1受体（NK1R）免疫标记的重叠，以及速激肽（[Sar9-Met（O2）11]-P物质）或[D-Ala2，N-MePhe4，Gly-ol5]-脑啡肽（DAMGO）分别增强或抑制呼吸频率，将该热点确定为preBötC。P2R激动剂的相对效力[2-甲硫基腺苷5'-二磷酸（2MeSADP）≈2-甲硫基腺苷5'-三磷酸（2MeSATP）≈腺苷5'-[γ-硫代]三磷酸四锂盐（ATPγS）≈ATP >>尿苷三磷酸（UTP）≈α，β-亚甲基腺苷5'-三磷酸（α，β-meATP）]以及MRS2179（2'-脱氧-N6-甲基腺苷-3'，5'-双磷酸）（P2Y1拮抗剂）对ATP反应的减弱表明，兴奋是由P2Y1受体介导的。ATP后的抑制作用（从未在对ATPγS的反应中观察到）取决于ATP的水解。这些数据在新生大鼠中证实，preBötC中的呼吸节律产生网络对P_{2Y1}R激活极为敏感，并提示P_{2Y1}R在呼吸运动控制中发挥作用，尤其是在缺氧期间发生的节律性P2R兴奋中。
