Ultrastructural detection of neuronally transported choleragenoid by postembedding immunocytochemistry in freeze-substituted Lowicryl HM20™ embedded tissue

Choleragenoid (cholera toxin B-fragment; CTB) is an anterograde, retrograde and transganglionic neuronal tracer. We describe a method for detecting CTB-labeled neuronal cell bodies, neurites and boutons at the ultrastructural level, using postembedding immunogold techniques on freeze-substituted Lowicryl HM20™ embedded nervous tissue. Primary afferents and motoneurons were labeled by injection of CTB in the dorsal ramus of the C2 spinal nerve of the rat. Following fixation with paraformaldehyde (4%) and glutaraldehyde (0.25%), tissue sections from the spinal cord C2 segment were freeze-substituted and embedded in Lowicryl HM20™ and subsequently processed with postembedding immunocytochemistry for CTB and glutamate. Immunogold particles indicating CTB immunoreactivity were found over primary afferents and motoneurons. In primary afferents in the central cervical nucleus (CCN) and motor nuclei, immunogold labeling was seen in boutons over vesicle-containing axoplasm and to a lesser extent over axoplasm devoid of vesicles, but not over mitochondria or axolemma. In motoneurons, immunogold particles were seen over the Golgi apparatus in the soma and over lysosomes in both soma and dendrites. Quantification of glutamate-like immunoreactivity in 20 CTB-labeled and 20 CTB-negative boutons in the neuropil was found similar, indicating that CTB does not interfere with the immunocytochemical detection of neuronal epitopes such as the transmitter substance glutamate.     

Extracellular characteristics of putative cholinergic neurons in the rat laterodorsal tegmental nucleus

The extracellular electrophysiological properties of neurons in the laterodorsal tegmental nucleus (LDT), a major source of cholinergic afferents to the thalamus, were studied in chloral hydrate-anesthetized rats. A combination of antidromic activation from the thalamus and histological verification of recording sites was used to correlate the identity of extracellular recordings in the rat LDT with cholinergic neurons in that region. All neurons antidromically activated by stimulation of the anteroventral thalamus were histologically verified to be within clusters of cholinergic (NADPH-d-positive) cells in the LDT or in the adjacent nucleus locus coeruleus (LC). The thalamically projecting LDT neurons had a homogeneous neurophysiological profile consisting of long duration action potentials (mean = 2.5 ms), slow conduction velocities (mean = 0.78 m/s), and lengthy chronaxie values (mean = 0.725 ms). The appearance and axonal characteristics of these neurons resembled those of noradrenergic LC neurons, but the two populations exhibited substantially different spontaneous activity patterns and sensory responsiveness. These characteristics may be useful in the preliminary identification of putative cholinergic neurons in vivo, and thereby provide a foundation for exploring the neuropharmacology, afferent modulation, sensory responsiveness and behavioral correlates of the brainstem cholinergic system.     

肿瘤抑制因子PTEN蛋白在自发性高血压大鼠主动脉平滑肌中的表达

目的：研究PTEN蛋白在雄性自发性高血压大鼠及SD大鼠主动脉中的表达。方法：取18周龄和26周龄雄性自发性高血压大鼠及同龄雄性SD大鼠主动脉石蜡包埋切片，免疫组织化学观察PTEN在主动脉平滑肌细胞中的表达。结果：SD大鼠主动脉平滑肌细胞中存在PTEN阳怀免疫反应产物。自发性高血压大鼠主动脉平滑肌细胞也存在PTEN阳性免疫反应产物，但免疫反应产物的染色强度明显低于SD大鼠。结论：SD大鼠主动脉平滑肌细胞PTEN高表达，自发性高血压大鼠主动脉平滑肌细胞PTEN表达呈下降趋势。提示PTEN可能在自发性高血压大鼠主动脉平滑肌细胞重塑过程中发挥作用。     

Endogenous vasopressin and baroreflex mechanisms

This article reviews the anatomical and functional evidence for ascending pathways from specific brain regions to the PVN and SON which could influence AVP release. The majority of evidence favours the main projection being from a region in the caudal VLM which may coincide with the noradrenergic neurons of the A₁ cell group. However, the transmitter(s) involved have yet to be identified, and whether the pathway is excitatory and/or inhibitory remains to be fully resolved.Anatomical and functional evidence is reviewed for descending projections from the SON and PVN to specific brain regions involved in cardiovascular control, and their possible involvement in baroreflex mechanisms is discussed. However, there is little unequivocal evidence that AVP is the main neurotransmitter utilized by descending projections from PVN to NTS and DMX. While, in some situations, circulating endogenous AVP exerts cardiovascular effects, details of its putative influences on baroreflex mechanisms are lacking.     

Facilitation of discrimination transfers under amphetamine: the relative control by S+ and S− and general transfer effects

Rats were trained in a Y-maze on a two-choice simultaneous black-white discrimination with either black or white as S⁺. Animals were then transferred to one of three discrimination tasks. In task 1 (New S^–), a new stimulus, either vertical or horizontal stripes, was substituted for the original S^–. In task 2 (New S⁺), a new stimulus, either vertical or horizontal stripes as in task 1, was substituted for the original S⁺. In task 3 (New S⁺/S^–) animals were trained on horizontal-vertical discrimination. The pre-trial administration of 1 mg/kg d-amphetamine facilitated the acquisition of the original black-white discrimination with both black as S⁺ and white as S⁺. Likewise, the drug improved performance in all three transfer conditions. However, the course of learning in the three transfer tasks was different in the placebo- and amphetamine-treated animals. Amphetamine-treated animals were disrupted more by a change in S⁺ than by a change in S^–, whereas the opposite pattern was evident in the placebo controls. When both discriminative stimuli were changed, placebo animals exhibited pronounced decrement in performance, whereas amphetamine animals exhibited excellent learning. The implications of these findings for the effects of amphetamine on discrimination learning are discussed.     

大鼠脑组织单胺类递质及其代谢产物的检测方法研究

目的 :研究大鼠脑组织中单胺类递质及其代谢产物的高效液相反相离子对色谱测定法。方法 :采用LiChrosorbC18,10 μm色谱柱 ,流动相为甲醇 :水 (4 0 :60 ) ,含 0 .0 2 8g LEDTANa2 ,0 .15g LSDS ,0 .2ml LH2 SO4(pH 2 .5～ 3 ) ,荧光检测波长 :λEX=2 85 ,λEM=3 3 3。结果 :对 87只大鼠脑组织中 4种单胺类递质及其代谢产物的含量进行了同时测定 ,高香草酸 (HVA) 2 .5 0～ 40 .0 μg ml、去甲肾上腺素 (NE) 0 .0 1～ 0 .5 0 μg ml、多巴胺 (DA)0 .0 5～ 1.0 0 μg ml、5 羟色胺 (5 HT) 0 .0 2 5～ 0 .5 0 μg ml,峰面积与其含量呈良好的线性关系。 结论 :该法操作简便、快速、准确 ,为组织中单胺类递质及其代谢产物检测的一种理想方法 ,并适用于临床相关研究。     

De-novo expression of vascular ecto-5′-nucleotidase and down-regulation of glomerular ecto-ATPase in experimental chronic renal transplant failure

Ischemic injury plays an important role in chronic renal transplant failure (CRTF). Down-regulation of ecto-adenosine triphosphatase (ATPase) in combination with up-regulation of ecto-5'-nucleotidase is a hallmark of ischemic injury. We studied the expression of renal ecto-5'-nucleotidase and ecto-ATPase in experimental renal transplantation. Fisher 344-to-Lewis allografted rats were either treated with an angiotensin-converting enzyme inhibitor (ACEi) or left untreated. Lewis-to-Lewis syngrafted rats served as controls. Untreated allografted rats developed proteinuria, glomerulosclerosis, and mild intimal hyperplasia. ACEi completely prevented focal and segmental glomerulosclerosis (FGS) and proteinuria, but significantly enhanced intimal hyperplasia. Untreated allografted rats revealed marked vascular ecto-5'-nucleotidase activity, which increased with ACEi. Vascular ecto-5'-nucleotidase activity was absent in syngrafted animals. Ecto-5'-nucleotidase activity correlated well with intimal hyperplasia. Glomerular ecto-ATPase expression was significantly reduced in untreated allografted rats compared to syngrafted rats and correlated well with the extent of FGS. ACEi prevented reduction in glomerular ecto-ATPase. We found de-novo expression of ecto-5'-nucleotidase at sites of renal intimal hyperplasia. Glomerular ecto-ATPase expression was markedly reduced in allografted rats and was prevented by ACEi. These enzyme expression patterns suggest local ischemic damage in experimental CRTF.     

正常小鼠胃肠运动昼夜节律及药物作用的影响

目的：观察小鼠胃肠运动昼夜节律及药物对节律的影响。方法：小鼠不同时间点灌胃给予蒸馏水及实验药物，同步测定胃排空及小肠推进运动情况。结果：小鼠胃排空及小肠推进运动存在昼夜节律；普瑞博思促进胃肠运动存在时间效应，但对节律无明显影响；阿托品抑制胃肠运动可使原有昼夜节律消失。结论：小鼠胃肠运动功能存在昼夜节律，药物对胃肠运动功能的作用存在时间效应，可使节律发生不同变化。     

Patterns of membrane potentials and distributions of the medullary respiratory neurons in the decerebrate rat

We analyzed the membrane potential of 161 respiratory neurons in the medulla of decerebrate rats which were paralyzed and ventilated. Three types of inspiratory (I) neurons were observed: those displaying progressive depolarization in inspiration (augmenting I neurons), those which gradually repolarized after maximal depolarization at the onset of inspiration (decrementing I neurons) and those exhibiting a plateau or bell-shaped membrane potential trajectory throughout inspiration (I-all neurons). Three types of expiratory (E) neurons were also encountered: those in which the membrane potential progressively depolarized (augmenting E neurons), those in which the membrane potential repolarized during the interval between phrenic bursts (decrementing E or post-I neurons) and those exhibiting a plateau or bell-shaped membrane potential trajectory throughout expiration (E-all neurons). Axonal projections of these medullary neurons were identified in the cranial nerves (n = 34), or in the spinal cord (n = 19) as revealed by antidromic stimulation and/or by reconstruction following horseradish peroxidase (HRP) labeling. The other 108 neurons were not antidromically activated (NAA) by the stimulations tested, or had their axons terminating inside the medulla as revealed by HRP labeling. All these respiratory neurons, except for 3 which were hypoglossal motoneurons, had their somata within the ventrolateral medulla, in the region of the nucleus ambiguus, homologous to the ventral respiratory group (VRG) of the cat. No dorsal respiratory group (DRG) was detected within the medulla of the rats. Due to this absence of a DRG, it is concluded that the neural organization of respiratory centers is quite different in cats and rats.     

不同情期大鼠血浆神经降压素含量的变化

本文试图阐明不同情期大鼠血浆神经降压素含量的变化。血浆神经降压素以无水乙醇提取后采用放射免疫分析法测定。间情期、动情前期和动情期大鼠血浆内神经降压素样免疫活性物质的含量依次分别为92.1±7.7ng/L,118.0±9.4ng/L 和144.3±10.2ng/L。动情期大鼠血浆神经降压素样免疲活性物质的含量较间情期大鼠有极显著差异(P<0.01)。