Effects of veratridine, tetrodotoxin and other drugs that alter electrical behaviour on secretion of melanocyte-stimulating hormone from melanotrophs of the pituitary pars intermedia

Since melanotrophs are electrically active and exhibit spontaneous Na spikes, a study was made of the effects, on melanotroph secretion, of drugs known to influence electrical properties. The output of melanocyte-stimulating hormone was measured from perifused neurointermediate lobes of mice or melanotrophs dispersed from such lobes of mice or rats. Veratridine (200 microM), which is known to increase Na permeability in a variety of cells, caused a large, although transient, increase in secretion from the melanotrophs that required extracellular Ca2+ and was blocked by the Na-channel blocker tetrodotoxin (1 microM). Tetraethylammonium (10 mM), which blocks K channels and thus prolongs the duration of the action potential in many cells, also stimulated secretion in the melanotrophs in a Ca-dependent manner. This response was not, however, blocked by tetrodotoxin, and is thus not attributable to prolongation of Na spikes in these cells. Moreover, tetrodotoxin did not inhibit basal secretion. The stimulant effect of veratridine on secretion in melanotrophs and its suppression by tetrodotoxin suggests that voltage-dependent Na channels can participate in the regulation of hormone output in these cells of the pituitary pars intermedia. However, the apparent lack of effect of tetrodotoxin on basal secretion suggests that the spontaneous Na spikes previously observed in these cells are not required for promoting the Ca influx which other evidence shows is important for basal secretion.     

Membrane properties of rat locus coeruleus neurones

Intracellular recordings were made from neurones in the locus coeruleus contained within a slice cut from rat pons and maintained in vitro. Most neurones fired action potentials spontaneously at frequencies of between 1 and 5 Hz; this did not arise from spontaneous synaptic input but appeared to result from endogenous properties of the membrane conductances. Under voltage clamp at potentials near threshold for action potential generation (? 55 mV) there was a persistent inward calcium current. This current became less with membrane hyperpolarization and was abolished at about ?70 mV. Two potassium currents were observed. The first had properties similar to that generally described as the “fast” potassium current (I_K,A); it flowed transiently (for about 200 ms) when the membrane potential passed from about ?65 to ?45 mV, and was blocked by 4-aminopyridine. The second was a calcium-activated potassium current (I_K,Ca); it flowed for several seconds following a burst of calcium action potentials. Spontaneous and evoked action potentials had both tetrodotoxin-sensitive and tetrodotoxin-resistant components. The latter was apparently due to calcium entry. The potential changes occurring during the spontaneous firing of locus coeruleus neurones could be reconstructed qualitatively from the ionic conductances observed. The membrane properties of the locus coeruleus neurones were remarkably uniform; however, about 5% of cells impaled within the region of the locus coeruleus were electrophysiologically distinct. These atypical cells had short duration action potentials, did not fire spontaneously and had large spontaneous depolarizing synaptic potentials.     

Electrical behaviour in a line of anterior pituitary cells (GH cells) and the influence of the hypothalamic peptide, thyrotrophin releasing factor

Anterior pituitary cells of the GH line, which secrete prolactin spontaneously, showed spontaneous action potential activity. Thyrotrophin releasing factor, which increases secretion in these cells, caused a prompt increase of action potential frequency. Potassium, another secretagogue, depolarized the cells and sometimes initiated a burst of action potentials at the onset of this effect. The action potentials persisted in tetrodotoxin-containing and Na-free media, but were suppressed by the Ca-channel blocker, methoxyverapamil. Moreover, elevating the extracellular Ca²⁺ concentration increased the amplitude of the action potentials. These action potentials therefore have a prominent Ca component. This endows them with a particular interest since secretory activity of these cells is known to be dependent on extracellular Ca²⁺. Ba²⁺, which can substitute for Ca²⁺ in maintaining secretion, also substituted for Ca²⁺ in the maintenance of the action potentials. In addition, Ba²⁺ prolonged action potentials remarkably: tetraethylammonium was less effective in this regard.The several parallels between known secretory behaviour and electrical phenomena encourage the view that analysis of electrical activity in anterior pituitary cells may provide useful clues to events involved in stimulus-secretion coupling and in the secretory control exerted by the brain.     

Different ion channel mechanisms between low concentrations of capsaicin and high concentrations of capsaicin and nicotine regarding peptide release from pulmonary afferents

Vagal nerve stimulation (1 Hz for 1 min), capsaicin (10^-8 M and 10^-6 M), resiniferatoxin (3 × 10^-10 M) and nicotine (10^-4 M) evoked a non-cholinergic bronchoconstriction in the isolated perfused guinea-pig lung preparation. Simultaneously there was an increase in the perfusate levels of calcitonin gene-related peptide-like immunoreactivity, suggesting release from sensory nerves. Both the bronchoconstriction and peptide release evoked by a low concentration of capsaicin (10^-8 M) and that evoked by nerve stimulation were depressed by tetrodotoxin, suggesting involvement of Na⁺ channel dependent depolarization. Since the effects of capsaicin (10^-8 M) and vagal nerve stimulation were inhibited by ω-conotoxin but not influenced by nifedipine, the Ca²⁺-channel involved is probably of N-type. Furthermore, the capsaicin analogue resiniferatoxin also evoked ω-conotoxin sensitive peptide release and bronchoconstriction. At the higher capsaicin concentration (10^-6 M), the functional response was only slightly inhibited by wconotoxin or tetrodotoxin indicating that capsaicin at this concentration evoked peptide release and functional effects through other mechanisms, probably involving Ca²⁺ fluxes in the non-selective cation channel associated with the proposed capsaicin receptor. The nicotine (10^-4 M) evoked peptide release and bronchoconstriction were only marginally influenced by ω-conotoxin or tetrodotoxin. It is concluded that the ion-channel mechanisms underlying the peptide releasing properties of antidromic nerve stimulation and low concentrations of capsaicin are similar and depend on action potential propagation, whereas capsaicin in high, toxic concentration and nicotine mainly act via receptor operated channels.     

Release of endogenous 5-hydroxytryptamine from the neural and the intermediate lobe of the rat pituitary gland evoked by electrical stimulation of the pituitary stalk

A calcium-dependent release of 5-hydroxytryptamine from the neural and intermediate lobe of the rat pituitary gland has been demonstrated following electrical stimulation of the pituitary stalk with stimulation parameters thought to evoke propagated action potentials. The 5-hydroxytryptamine release from the intermediate lobe was double that from the neural lobe. The mass of the intermediate lobe of the rat is about 80% of that of the neural lobe [Holzbauer, Racké, Mann, Cooper, Cohen, Krause and Sharman (1984) J. Neural Transm. 59, 91-104]. The relatively high overflow of 5-hydroxytryptamine from the intermediate lobe agrees with immunohistochemical studies in which a larger number of 5-hydroxytryptamine fibres were seen in the intermediate lobe than in the neural lobe. The present results have demonstrated that the rat hypophysis contains neuronal 5-hydroxytryptamine. They also suggest that this amine may act as a neurotransmitter substance in the neural and intermediate lobe.     

Effects of benactyzine on action potentials and contractile force of guinea pig papillary muscles

ＩＮＴＲＯＤＵＣＴＩＯＮＢｅｎａｃｔｙｚｉｎｅｉｓａｓｙｎｔｈｅｔｉｃｂｌｏｃｋｅｒｏｆＭ ｒｅｃｅｐｔｏｒｐｏｓ ｓｅｓｓｉｎｇｓｍｏｏｔｈｍｕｓｃｌｅｒｅｌａｘａｎｔｐｒｏｐｅｒｔｉｅｓａｎｄｑｕｉｎｉｄｉｎｅ ｌｉｋｅａｃｔｉｏｎｏｎｈｅａｒｔ[1] .Ｂｅｎａｃｔｙｚｉｎｅｉｓｅｆｆｅｃｔｉｖｅｉｎｒｅｓｔｏｒ ｉｎｇｃｉｒｃｕｌａｔｏｒｙｆｕｎｃｔｉｏｎｄｉｓｔｕｒｂｅｄｂｙｏｒｇａｎｏｐｈｏｓｐｈａｔｅｐｏｉ ｓｏｎｉｎｇ[2 ,3] .Ｐｒｅｖｉｏｕｓｓｔｕｄｉｅｓｓｕｇｇｅｓｔｅｄｔｈａｔｂｅｎａ…     

河豚毒素与局麻药联合应用对家兔齿髓刺激所致疼痛的对抗作用

河豚毒素(tetrodotoxin，TTX)是一种毒性极高的非蛋白神经毒素。具有很强的局麻作用。其效力比目前常用局麻药强万倍以上。持续时间长。将TTX与常用局麻药联合应用于齿髓镇痛。国内外文献尚未报道。我们设想将极微量TTX与常用局麻药利多卡因、丁卡因联合给药，通过齿髓刺激试验，测定其局部镇痛作用，观察是否具有协同或增强局麻效果，以探讨TTX作为协同局麻药应用于临床的可能。     

家兔眼球后注射替曲朵辛的麻醉效果及毒副作用研究

目的 对替曲朵辛(TTX)溶液球后注射的麻醉效应、安全性和视网膜毒副作用进行初步评价。方法 健康成年家兔随机分组，每组6只，其中有眼球后注射不同剂量TTX及利多卡因溶液，左眼给予等量溶剂对照(柠檬酸钠缓冲液，pH4．3)；分别采用角膜感觉试验、裂隙灯、视网膜电图、光电镜检查进行观察。结果 球后注射10μg TTX可致家兔全部死亡；≤5μg剂量家兔全部存活；具有明显的局部麻醉作用；5μg剂量组可观察到轻微视网膜毒性作用；而≤2．5μg则未见眼部毒性反应。结论 球后注射低剂量TTX对家兔角膜可产生长效局部麻醉作用且无明显眼毒性反应，有希望应用于治疗某些疼痛性眼病及术中镇痛。     

河豚鱼干中毒的动物实验研究及在法医鉴定中的应用

目的:研究河豚鱼干中的毒性成分对小白鼠肝、肾等脏器的损害。方法:应用乙酸浸泡法提取涉案河豚鱼干中的含毒浸出液,在1h、2h、3h、4h和6h共5个不同时间段,对小白鼠采用灌胃和腹腔注射方法,制造实验中毒模型。结果:实验3h、4h组损伤最明显,死亡率达100%;河豚鱼毒素(Tetrodotoxin,TTX)导致实验小白鼠的肝、肾组织细胞中毒坏死,所有实验动物均出现呼吸急促、步态不稳和口尾紫绀等TTX中毒症状。结论:实验证实检案确系河豚鱼干中毒。     

Munc13 genotype regulates secretory amyloid precursor protein processing via postsynaptic glutamate receptors

The amyloid precursor protein (APP) can be proteolytically degraded via non-amyloidogenic α-secretase and amyloidogenic β-secretase pathways. Previously, we have identified the presynaptic protein Munc13-1 as a diacylglycerol/phorbolester (DAG/PE) receptor that contributes to secretory, non-amyloidogenic APP processing after PE stimulation. Here, we used organotypic brain slice cultures from wild-type mice and from Munc13-1 knock-out (KO), Munc13-2 KO and Munc13-1/2 double KO (DKO) mice for pharmacological stimulation experiments. First, we demonstrate that neuronal populations and synaptic components important for secretory APP processing develop normally in organotypic brain slice cultures of all genotypes analyzed. Blockade of voltage-gated Na⁺ channels by tetrodotoxin reduced the PE-stimulated secretory APP processing, whereas depolarization by high extracellular K⁺ concentration evoked APP secretion. Additionally, the PE-stimulated APP secretion from Munc13-1 KO brain slices was significantly lower than that from wild-type brain slices. This effect was not observed in brain slices from Munc13-2 KO mice, which is consistent with the lower abundance and subpopulation-specific distribution of Munc13-2 in presynaptic elements. In Munc13-1/2 DKO brain slices, the deficiency of Munc13-1 dominated the effect of APP processing. The Munc13-1 KO effect on APP processing could be rescued by the stimulation of postsynaptic glutamatergic receptors. This indicates that lack of postsynaptic glutamate receptor stimulation in Munc13-1 KO brain slice cultures but not presynaptic mechanisms account for compromised APP processing. We conclude that organotypic brain slices cultures are a valuable tool for studying APP processing pathways in intact neuronal circuits and that neuronal activity is important for maintenance of the non-amyloidogenic APP processing.