Nonlinear analysis of intracellular recording of rostral ventrolateral medulla neurons using whole-cell patch-clamp technique

为了确定血压升高的机制，我们比较了新生Wistar-Kyoto （WKY）和自发性高血压大鼠（SHR）延髓吻侧腹外侧（RVLM）神经元的电生理特征。利用全细胞膜片钳技术，我们在保留交感神经网络的脑干-脊髓制备中检测了RVLM神经元的特性。SHR组（n=56） RVLM神经元的基线膜电位较WKY组（n=33）浅（-49±1 mV比-53±2 mV），放电速度快（5.1±0.3 Hz比4.1±0.5 Hz）。血管紧张素II（6微mol/L）在SHR组（8.9±1.9 mV）比WKY组（1.8±0.8 mV）对RVLM神经元的去极化作用更大。血管紧张素II型1 （AT_1）受体拮抗剂坎地沙坦（0.12微mol/L）诱导SHR RVLM神经元的膜超极化（4.9±1.1 mV）和放电速率明显降低，而WKY RVLM神经元的放电速率无明显降低。与AT_2受体拮抗剂PD123319 （60 micromol/L）融合后，超极化消失，表明血管紧张素II通过AT_2受体参与了这种超极化。RVLM神经元的电生理特性及其对血管紧张素II的反应在WKY和SHR之间存在差异。坎地沙坦引起的明显的超极化和放电速率降低提示内源性血管紧张素II实际上与RVLM神经元上的AT_1受体结合，从而促进高血压大鼠膜电位升高和放电速率加快。RVLM神经元的这些差异暗示了一种可能导致血压升高的机制。此外，通过记录清醒WKY和SHR时的血压（BP）、心率、肾交感神经活动（RSNA）和同侧肾血流量（RBF），我们发现RSNA和BP之间以及RSNA和RBF之间的传递函数在0.05和0.80 Hz时出现一致性峰值，低于呼吸和心脏相关波动。SHR的相干性明显高于WKY，增益明显小于WKY。互信息值显示，在0 ~ 0.1 Hz范围内，RSNA与BP和RBF均存在较强的线性和非线性关系，且在2个菌株中关系相似。高相干性表明高血压大鼠BP和RBF与RSNA的线性相关性更强，而正常大鼠的非线性相关性更强。时滞值表明，在闭环条件下，RSNA先于BP和RBF。在有意识的大鼠中，中枢神经系统产生的明显的低频RSNA振荡产生血流动力学波动，独立于气压反射。少

To determine mechanisms underlying blood pressure elevation, we compared electrophysiological characteristics of neurons in the rostral ventrolateral medulla (RVLM) of neonatal Wistar-Kyoto (WKY) and spontaneously hypertensive rats (SHR). Using the whole-cell patch-clamp technique, we examined the properties of RVLM neurons in brainstem-spinal cord preparations with a preserved sympathetic neuronal network. The baseline membrane potential of RVLM neurons was shallower (-49±1 mV vs.-53±2 mV) and the firing rate was faster (5.1±0.3 Hz vs.4.1±0.5 Hz) in SHR (n=56) than in WKY (n=33). Superfusion with angiotensin II (6 micromol/L) depolarized the RVLM neurons in SHR (8.9±1.9 mV) to a greater extent than in WKY (1.8±0.8 mV). Angiotensin II type 1 (AT_1) receptor antagonist, candesartan (0.12 micromol/L), induced significant membrane hyperpolarization (4.9±1.1 mV) and a decrease in the firing rate in RVLM neurons of SHR but not in those of WKY.After superfusion with the AT_2 receptor antagon … More ist, PD123319 (60 micromol/L), the hyperpolarization was abolished, suggesting angiotensin II action through AT_2 receptors to be involved in this hyperpolarization. Electrophysiological properties of RVLM neurons and their response to angiotensin II differ between WKY and SHR.The significant hyperpolarization and decreased firing rate caused by candesartan suggest that endogenously generated angiotensin II actually binds to AT_1 receptors on RVLM neurons, thus tonically contributing to a higher membrane potential and a faster firing rate in hypertensive rats. These differences in RVLM neurons suggest a mechanism possibly leading to elevation in blood pressure.Additionally, by recording blood pressure (BP), heart rate, renal sympathetic nerve activity (RSNA), and ipsilateral renal blood flow (RBF) in conscious WKY and SHR, we found coherence peaks of the transfer function between RSNA and BP and between RSNA and RBF at 0.05 and 0.80 Hz, below the respiratory-and cardiac-related fluctuations. The coherence was significantly higher and gain smaller in SHR than in WKY.Mutual information values showed strong linear and nonlinear relationships between RSNA and both BP and RBF at 0-0.1 Hz, and the relationships were similar in the two strains. The higher coherence implies that BP and RBF were more linearly correlated with RSNA in hypertensive rats, whereas the nonlinear correlation was greater in normotensive rats. Time delay values demonstrated that RSNA preceded BP and RBF under closed-loop conditions. Distinct low-frequency RSNA oscillations arising from the central nervous system generated hemodynamic fluctuations in conscious rats, independently of the baroreflex. Less