Huang R Q, Erlichman J S, Dean J B
Department of Physiology and Biophysics, Wright State University, School of Medicine, Dayton, OH 45435, USA.
Neuroscience. 1997 Sep;80(1):41-57. doi: 10.1016/s0306-4522(97)00017-1.
Anatomically coupled neurons (17 of 137) and non-coupled neurons (120 of 137), in and near the nucleus tractus solitarius and dorsal motor nucleus (i.e. solitary complex), were studied by rapid perforated patch recording in slices (rat, 150-350 microm thick, postnatal day 0-21) before, during and after exposure to hypercapnic acidosis. Anatomical coupling refers to the intercellular transfer of Lucifer Yellow and Biocytin into adjoining neurons, presumably via gap junctions [see Dean et al. (1997) Neuroscience 80, 21-40]. Eighty-six per cent of the anatomically coupled neurons (12 of 14) were depolarized by hypercapnic acidosis, a response referred to as CO2 excitation or CO2 chemosensitivity. In all, 28% (12 of 43) of the CO2-excited neurons were anatomically coupled to at least one other neuron. None (0 of 39) of the CO2-inhibited neurons were anatomically coupled, and only 4% (two of 46) of the CO2-insensitive neurons were anatomically coupled. Increasing the fractional concentration of CO2 from five to 10 and 15% in constant bicarbonate (26 mM) decreased intracellular pH (control 7.3 7.4, 22-25 degrees C) by approximately 1.0 and 1.5 pH units, respectively, as measured using the pH-sensitive fluorescent dye, 2',7'-bis (2-carboxyethyl)-5,6-carboxyfluorescein. Nine of the anatomically coupled neurons (six CO2-excited, one CO2-insensitive and two unidentified) exhibited spontaneous electrotonic postsynaptic potential-like activity, suggesting that they were also electrotonically coupled. During hypercapnic acidosis, the amplitudes of electrotonic postsynaptic potentials were unchanged, concomitant with small changes in input resistance. The frequency of electrotonic postsynaptic potentials increased during hypercapnic acidosis in many anatomically coupled neurons (eight of nine), indicating that both neurons of the coupled pair were stimulated. Cell-cell coupling occurred preferentially in and between CO2-excited neurons of the solitary complex. Further, CO2-excited neurons were not electrotonically uncoupled during intracellular acidosis, in contrast to the effect that decreased intracellular pH has on many other types of coupled cells. It was not determined whether anatomical coupling was affected by hypercapnic acidosis since dye mixture was always administered under normocapnic conditions. The high correlation between anatomical coupling, electrotonic coupling activity and CO2-induced depolarization suggests that cell-cell coupling is an important electroanatomical feature in CO2-excited neurons of the solitary complex. CO2-excited neurons have been hypothesized to function in central chemoreception for the cardiorespiratory control systems, suggesting that cell cell coupling may contribute in part to central chemoreception of CO2 and H+.
采用快速穿孔膜片钳记录技术，在离体脑片（大鼠，出生后0 - 21天，脑片厚度150 - 350微米）中，于暴露于高碳酸血症性酸中毒之前、期间和之后，对孤束核和背运动核（即孤束复合体）及其附近的解剖学耦联神经元（137个中有17个）和非耦联神经元（137个中有120个）进行了研究。解剖学耦联是指荧光黄和生物胞素大概通过缝隙连接向相邻神经元的细胞间转移[见Dean等人（1997年），《神经科学》80卷，21 - 40页]。86%的解剖学耦联神经元（14个中有12个）在高碳酸血症性酸中毒时发生去极化，这种反应被称为CO₂兴奋或CO₂化学敏感性。总共有28%（43个中有12个）的CO₂兴奋神经元与至少一个其他神经元存在解剖学耦联。CO₂抑制神经元中无一（39个中有0个）存在解剖学耦联，CO₂不敏感神经元中只有4%（46个中有2个）存在解剖学耦联。在固定碳酸氢盐（26 mM）条件下，将CO₂的分数浓度从5%增加到10%和15%，使用pH敏感荧光染料2',7'-双（2 - 羧乙基）-5,6 - 羧基荧光素测量，细胞内pH（对照7.3 - 7.4，22 - 25℃）分别降低约1.0和1.5个pH单位。9个解剖学耦联神经元（6个CO₂兴奋、1个CO₂不敏感和2个未明确类型）表现出自发性电紧张性突触后电位样活动，表明它们也存在电耦联。在高碳酸血症性酸中毒期间，电紧张性突触后电位的幅度不变，同时输入电阻有小的变化。在许多解剖学耦联神经元（9个中有8个）中，高碳酸血症性酸中毒期间电紧张性突触后电位的频率增加，表明耦联对中的两个神经元都受到刺激。细胞间耦联优先发生在孤束复合体的CO₂兴奋神经元内部和之间。此外，与细胞内pH降低对许多其他类型耦联细胞的影响相反，CO₂兴奋神经元在细胞内酸中毒期间并未发生电脱耦联。由于染料混合物总是在正常碳酸血症条件下给药，所以未确定解剖学耦联是否受高碳酸血症性酸中毒影响。解剖学耦联、电耦联活动与CO₂诱导的去极化之间的高度相关性表明，细胞间耦联是孤束复合体中CO₂兴奋神经元的一个重要电解剖学特征。据推测，CO₂兴奋神经元在心肺控制系统的中枢化学感受中起作用，这表明细胞间耦联可能部分有助于对CO₂和H⁺的中枢化学感受。
