Quantitative gap junction alterations in mammalian heart cells quickly frozen or chemically fixed after electrical uncoupling

Summary The gap junction morphology was quantified in freeze-fracture replicas prepared from rat auricles that had been either quickly frozen at 6 K or chemically fixed by glutaraldehyde, in a state of normal cell-to-cell conduction or in a state of electrical uncoupling. The general appearance of the gap junctions was similar after both preparative procedures. A quantitative analysis of three gap junctional dimensions provided the following measurements in the quickly frozen conducting auricles (mean±sd): (a) P-face particles' diameter 8.27±0.74 nm (n =5709), (b) P-face particles' center-to-center distance 10.78±2.12 nm (n=4800), and (c) E-face pits' distance 9.99±2.19 nm (n=1600). Corresponding values obtained from chemically fixed tissues were decreased by about 3% for the particle's diameter and about 5% for the particles' and pits' distances. Electrical uncoupling by the action of either 1 mM 2–4-dinitrophenol (DNP), or 3.5 mMn-Heptan-1-ol (heptanol), induced a decrease of the particle's diameter, which amounted to –0.69±0.01 nm (mean ±se) in the quickly frozen preparations and –0.71±0.01 nm in the chemically fixed ones. The particles' distance was decreased by –0.96±0.04 nm in the quickly frozen samples and by –0.90 ±0.03 nm in the chemically fixed ones and the E-face pits' distance was similarly reduced. All differences were statistically significant (P<0.001 for all dimensions). Electrical recoupling after the heptanol effect promoted a return of these gap junctional dimensions towards normal values, which was about 50% complete within 20 min. It is concluded that very similar morphological alterations of the gap junctional structure are induced in the mammalian heart by different treatments promoting electrical uncoupling and that these conformational changes appear independently of the preparative procedure. The suggestion that the observed decrease of the particles' diameter is genuinely related to the closing mechanism of the unit cell-to-cell channel set in thei centers is thus confirmed.     

电离辐射引起胃排空延迟机理的初步探讨

本文观察了800rad照射后胃肠肌电的变化及切斷内脏神经与电刺激迷走神经对官的影响,初步分析了电离辐射引起胃排空延迟的机理。结果是:800rad照后大鼠胃肠肌电的峰波平均振幅胃幽部窦部照后1小时至第4天明显降低(P<0.05);幽门括约肌政变不大;十二指肠球部照后第3～7天明显下降(P<0.01)。切断内脏神经后照射大鼠胃肠肌电的峰波平均振幅胃幽门窦部变化不大,幽门括约肌照后1小时至第5天明显升高(P<0.05),十二指肠球部照后第3～7天明显下降(P<0.05)。800rad照后第4天大鼠胃对电刺激迷走神经的反应性明显降低(P<0.01)。上述结果表明:照后胃排空延迟的原因不仅与神经-体液调节紊乱有关,而且与射线损伤了平滑肌,使其兴奋性暂时的功能性降低有关。此外照后由于十二指肠球部运动的变化而使内感受器对胃运动的调节发生紊乱也是值得重视的因素。     

延髓中缝核区注射谷氨酸钠降低大鼠动脉血压的作用

本工作研究了大鼠延髓中缝核在心血管活动调整中的作用。通过埋植套管向大鼠延髓中缝核区注射神经元兴奋剂谷氨酸钠5-μg可明显降低血压,抑制交感神经传出活动,心率无显著改变。降血压效应可被脊髓蛛网膜下腔注射5-HT受体阻断剂肉桂硫胺50μg,或腹腔注射赛庚啶15mg/kg所部分对抗;并可被皮下注射阿片受体阻断剂纳洛酮1mc/kg所削弱。结果提示:延髓中缝核-脊髓5-HT下行通路对脊髓的血管调节功能起抑制性影响,内源性阿片样物质可能参与该机制。     

Molecular motion underlying activation and inactivation of sodium channels in squid giant axons

Summary Measurements of the changes in birefringence associated with changes in membrane potential were made with internally perfused squid giant axons in low sodium solutions at 0–8°C. The time course of the birefringence changes share many properties of the gating (polarization) currents previously studied in this nerve. Both can be demonstrated as an asymmetry in the response to voltage pulses symmetrical about the resting potential which is not present about a hyperpolarized holding potential. Both have a rapid relaxation, which precedes the sodium permeability change. Both exhibit an initial delay or rising phase. Both are reversibly blocked by perfusion with 30mm colchicine; neither are altered by changes on sodium concentrations or 300nm tetrodotoxin. The birefringence response has a decrease in the amplitude of the rapid relaxation associated with the appearance of a slow relaxation. This is similar to the immobilization of fast gating charges which parallels sodium current inactivation.The amplitude of the birefringence and the gating current responses is consistent with a change in the alignment of several hundred peptide bonds per sodium channel.     

Cholinesterases in Single Nerve Cells Isolated from the Locus Ceruleus and from Nucleus of the Facial Nerve of the Rat: A Microgasometric Study

Cholinesterase activity in single nerve cell bodies isolated from the locus ceruleus and nucleus of the facial nerve of the rat was analyzed by the microgasometric method. Acetylcholinesterase activity is about the same in both types of cells. Nonspecific cholinesterase is present in noradrenergic cells of the locus ceruleus but not in the cholinergic cells of the nucleus of the facial nerve. The total activity of cholinesterases and the activity of acetylcholinesterase in nerve cell bodies isolated from the locus ceruleus remains practically unchanged from the tenth postnatal day until the age of 24 months. Depletion of noradrenaline by a high dose of reserpine does not influence the total activity of cholinesterases in nerve cell bodies of locus ceruleus.     

Changes in Protein Content of Goldfish Optic Nerve During Degeneration and Regeneration Following Nerve Crush

Abstract: After the goldfish optic nerve was crushed, the total amount of protein in the nerve decreased by about 45% within 1 week as the axons degenerated, began to recover between 2 and 5 weeks as axonal regeneration occurred, and had returned to nearly normal by 12 weeks. Corresponding changes in the relative amounts of some individual proteins were investigated by separating the proteins by two-dimensional gel electrophoresis and performing a quantitative analysis of the Coomassie Brilliant Blue staining patterns of the gels. In addition, labelling patterns showing incorporation of [³H]proline into individual proteins were examined to differentiate between locally synthesized proteins (presumably produced mainly by the glial cells) and axonal proteins carried by fast or slow axonal transport. Some prominent nerve proteins, ON1 and ON2 (50–55 kD, pI ～6), decreased to almost undetectable levels and then reappeared with a time course corresponding to the changes in total protein content of the nerve. Similar changes were seen in a protein we have designated NF (～130 kD, pI ～5.2). These three proteins, which were labelled in association with slow axonal transport, may be neurofilament constituents. Large decreases following optic nerve crush were also seen in the relative amounts of α- and β-tubulin, which suggests that they are localized mainly in the optic axons rather than the glial cells. Another group of proteins, W2, W3, and W4 (35–45 kD, pI 6.5–7.0), which showed a somewhat slower time course of disappearance and were intensely labelled in the local synthesis pattern, may be associated with myelin. A small number of proteins increased in relative amount following nerve crush. These included some, P1 and P2 (35–40 kD, pIs 6.1–6.2) and NT (～50 kD, pI ～5.5), that appeared to be synthesized by the glial cells. Increases were also seen in one axonal protein, B (～45 kD, pI ～4.5), that is carried by fast axonal transport, as well as in two axonal proteins, HA1 and HA2 (～60 and 65 kD respectively, pIs 4.5–5.0), that are carried mainly by slow axonal transport. Other proteins, including actin, that showed no net changes in relative amount (but presumably changed in absolute amount in direct proportion to the changes in total protein content of the nerve), are apparently distributed in both the neuronal and nonneuronal compartments of the nerve.     

Decreased Incorporation of [3H]Inositol and [3H]Glycerol into Glycerolipids of Sciatic Nerve from the Streptozotocin Diabetic Rat

The incorporation of [3H]myo-inositol into individual phosphoinositides and of [3H]glycerol into glycerolipids was determined in sciatic nerve obtained from normal and streptozotocin diabetic rats and incubated in vitro. The uptake of inositol into lipid was approximately linear with time. More than 80% of the label was present in phosphatidylinositol with the remainder divided about equally between phosphatidylinositol phosphate and phosphatidylinositol-4,5-bisphosphate. Labeling was unchanged 2 weeks after induction of diabetes, but was reduced by 32% after 20 weeks of the disease. Glycerol incorporation occurred primarily into phosphatidylcholine and triacylglycerol and was depressed up to 45% into major phosphoglycerides in nerves from both 2- and 20-week diabetic animals. Triacylglycerol labeling was also substantially decreased, and the reduction was comparable in intact and epineurium free nerve, suggesting that a metabolically active pool of this compound, which is sensitive to hyperglycemia and/or insulin deficiency, is located in or immediately adjacent to the nerve fibers. The considerable decline in incorporation of these lipid precursors in diabetic nerve may be related to impaired inositol transport and to decrease overall energy utilization by the tissue.     

Effect of calcitonin gene-related peptide on the neuroeffector mechanism of sympathetic nerve terminals in rat vas deferens

In order to evaluate the mode of action of calcitonin gene-related peptide (CGRP) on the neuroeffector mechanism of peripheral sympathetic nerve fibers, the effects of CGRP were tested on the electrical stimulated and the non-stimulated preparations of the isolated rat vas deferens. The contractile responses, which were mediated predominantly by activation of postganglionic noradrenergic nerve fibers, were dose-dependently inhibited by CGRP in concentrations ranging from 0.1 to 10 nM. The inhibitory response produced by CGRP in high concentrations (greater than 2 nM) usually returned to the control level at 20-30 min and were rarely tachyphylactic. The inhibitory action of CGRP was not modified by pretreatment with 10(-7) M propranolol or 10(-7) M atropine. Contractions produced by exogenous norepinephrine (NE) and 5-hydroxytryptamine (5-HT) in unstimulated preparations were not affected by pretreatment with CGRP in a low concentration (less than 2 nM). On the other hand, the contractions were slightly reduced 1 min after pretreatment with CGRP in high concentrations (greater than 5 nM), which recovered in 15 min after constant flow washout. High concentrations of CGRP also caused a concentration-dependent relaxation on the precontracted preparations produced by high potassium (60 mM K+) solution. These results suggest that CGRP in high concentrations (greater than 5 nM) may have a non-specific inhibitory action on the postsynaptic plasma membrane of the smooth muscle cell and a postulated CGRP receptor exists presynaptically in the rat vas deferens and that CGRP may inhibit the release of NE during adrenergic nerve stimulation.     

刺激家兔颈交感神经对颈动脉窦反射的影响

在36只麻醉家兔观察了电刺激颈交感神经(CSN)对颈动脉窦压力感受器(CSB)活动的影响。所得结果如下:(1)电刺激 CSN 可使夹闭颈动脉引起的加压反射消失或倒转,△BP 从刺激前的 39.5±3.6mmHg 变为刺激时的-0.31±5.4mmHg(P<0.001)。(2)在电刺激CSN 时,静注新福林所诱发的颈动脉窦压力感受器-心率反射增强,表现为反射性心率减慢较刺激前更为明显。(3)在以50—200mmHg 的压力充胀两侧颈动脉窦的条件下,刺激 CSN 引起窦内压与平均动脉压的关系曲线下移,与刺激前曲线相比有明显差异(P<0.01)。(4)切断 CSN 后,动脉血压有所升高,提示 CSN 对 CSB 活动有紧张性调节作用。以上结果比较明确地表明家兔 CSN 对 CSB 活动有调节作用。此作用可能是 CSN 作用于窦壁平滑肌而间接引起的。     

电针和吗啡对伤害性刺激猫内脏大神经引起的海马诱发电位的影响

在氯醛糖麻醉并箭毒化的50只猫中,观察了强电流刺激内脏大神经时的背侧海马诱发电位(Hippocampal evoked potential,HEP)以及电针和吗啡对其影响。结果显示,用20V电压刺激内脏大神经可以兴奋包括 Aδ和 C 纤维在内的几乎全部纤维,并且可以在海马内记录到波形稳定的 HEP。以负波为主的 HEP 在海马锥体细胞层附近以及下脚复合体的表浅层记录到,以正波为主的 HEP 在海马锥体细胞顶树突的中下部记录明显。只有刺激强度达到兴奋内脏大神经的 Aδ纤维时才可记录出 HEP。吗啡对 HEP 有显著的抑制作用。电针也可以明显抑制 HEP 的振幅,并有后作用。上述结果提示,内脏大神经中的感觉纤维,包括痛纤维的冲动可以传至海马。