ATP-Citrate Lyase and Other Enzymes of Acetyl-CoA Metabolism in Fractions of Small and Large Synaptosomes from Rat Brain Hippocampus and Cerebellum

The activities of choline acetyltransferase and ATP-citrate lyase were significantly correlated (r = 0.995) in fractions of small and large synaptosomes isolated from rat hippocampus and cerebellum. The activities of these two enzymes did not correlate with those of pyruvate dehydrogenase, carnitine acetyltransferase, citrate synthase, acetyl-CoA synthetase, lactate dehydrogenase, or with the rate of high-affinity glutamate uptake in the synaptosomal fractions. The results provide additional evidence linking ATP-citrate lyase to the cholinergic system in the brain.     

Lack of Effect of Repeated Electroconvulsive Shock on [3H]Spiroperidol and [3H]5-Hydroxytryptamine Binding and Cholinergic Parameters in Rat Brain

: Repeated electroconvulsive shock (ECS) administered on alternate days for 10 days produced no changes in rat striatal [³H]spiroperidol binding measured 24 h after the last shock compared to anaesthetised controls. Similarly, there was no change in whole brain specific [³H]5-HT binding. Sodiumdependent high affinity [³H]choline uptake (HAUC) and ChAT were also unaltered in striatal and hippocampal samples following repeated ECS. Acute administration of Pentylenetetrazol did produce an increase in hippocampal HAUC immediately postictally. However, ECS (XI) did not change HAUC measured 1 h postictally. An effect of halothane on HAUC was noted in these experiments indicating the importance of an evaluation of anaesthetic effects in ECS studies.     

Fine structure and innervation of the avian adrenal gland

Summary According to their ultrastructure and histochemistry three types of efferent nerve fibers can be distinguished in the bird's adrenal gland. The main part is made up of cholinergic fibers recognizable by a positive reaction for acetylcholinesterase and two specific populations of granules within the synaptic ending. Synaptic vesicles measuring 300 to 500 Å in diameter and dense-cored vesicles with a diameter of about 1 000 Å are discernible.In the periphery of the gland cholinergic axons for the innervation of adrenal cells form large bundles surrounded by a perineural sheath. The bundles cross the capsule and are situated within the adrenal chromaffin cords or at their periphery. Finally small groups of fibers enter a group of chromaffin cells which are surrounded by a basal lamina and which consist of about a dozen or more cells producing adrenaline and noradrenaline. Synaptic endings occur, above all in passeriform species, in the center of a chromaffin cell complex. They are either attached to the innervated cells or their dendrite-like processes, or embedded into the cells, or connected to short spines of the innervated cells. Synaptic and dense-cored vesicles leave the bouton by exocytosis. One synaptic terminal may innervate up to three A- or NA-cells. The existence of different types of synapses for A- and NA-cells cannot be excluded.Supported by a grant from the Deutsche Forschungsgemeinschaft (Un 34/1).     

Ultrastructural distribution of alpha-bungarotoxin binding sites in the suprachiasmatic nucleus of the rat hypothalamus

Summary The distribution of (¹²⁵I) alpha bungarotoxin (-BTX) binding sites in the suprachiasmatic nucleus (SCN) of the adult female rat was examined by electron-microscopic autoradiography. The ultrastructural distribution of silver grains was analysed by line source, direct point count, and 50% probability circle methods. Real grain distribution was significantly different from that of randomly generated hypothetical grains. Line source analysis demonstrated two populations of sources: one associated with membranes, and one inside neuronal structures. Probability circle analysis of shared grains indicated that membrane-bound-radioactive sources were mainly asssociated with axo-dendritic appositions. Only a small proportion of labeled neuronal interfaces exhibited synaptic differentiations in the plane of section. However, the compartment containing synaptic terminals was the most enriched when comparing real to hypothetical grains. Probability circle analysis of exclusive grains demonstrated that sources that were not associated with neuronal plasma membranes were likely to be within nerve cell bodies and dendrites. It is concluded that the majority of specifically labeled -BTX binding sites in the SCN is membrane bound, and may be associated with axodendritic synaptic transmission. The presence of a significant proportion of the label in the soma and dendrites of suprachiasmatic neurons 24 h after ventricular infusion suggests that some of the labeled binding sites (junctional or nonjunctional) may be internalized within these two compartments.     

Actions of amantadine at synaptic and extrasynaptic cholinergic receptors in the central nervous system of the cockroach Periplaneta americana

The effects of amantadine were investigated on cercal afferent, giant interneurone synapses and on the cell body membrane of the fast coxal depressor motoneurone (D_f), in the cockroach Periplaneta americana. Bath-applied amantadine at concentrations above 2.0 × 10^?5 M significantly reduced the amplitude of unitary and compound epsps recorded by sucrose-gap methods from cercal afferent, giant interneurone synapses in the desheathed sixth abdominal ganglion. Complete block of synaptic transmission was achieved at 1.0 × 10^?3 M amantadine. Synaptic blockade, which was not accompanied by changes in resting potential, was almost fully reversed by washing the ganglion in normal saline. From the dose-dependence of the synaptic blocking action, a Hill coefficient of 0.94 was estimated, indicating that there is no co-operativity in the binding of amantadine to its site of action.Bath-application of amantadine (5.0 × 10^?5 M) resulted in a parallel shift to the right of the dose-response curve for the depolarizing postsynaptic actions of acetylcholine. Nevertheless, even at a concentration of 2.0 × 10^?3 M, amantadine failed to protect the synaptic acetylcholine receptor/ion channel complex from the blocking action of α-bungarotoxin (5.0 × 10^?7 M). In addition, the block by amantadine of the acetylcholine-induced current recorded from the cell body membrane of the fast coxal depressor motoneurone (D_f), was strongly dependent on membrane potential in the range ? 120mV to ? 70mV. An action of amantadine at the open acetylcholine receptor/ion channel complex is proposed.     

‘Neurosecretion’ by a classic cholinergic innervation apparatus

Summary Nerve terminals forming typical synapses with adrenal chromaffin tissues have been examined in the goldfish, frog (Rana pipiens), hamster and rat. Presumptive secretory inclusions present in the terminals are of two distinct types. Electron-lucent synaptic vesicles 30–50 nm in diameter are densely clustered adjacent to membrane thickenings and presumably discharge their contents into the synaptic clefts. Secretory granules (i.e. large dense-cored vesicles) 60–100 nm in diameter are more abundant in other parts of the terminals. Sites of granule exocytosis have been observed in each of the animals investigated. They are usually encountered within apparently undifferentiated areas of plasmalemma and only rarely occur within synaptic thickenings. Granule exocytosis from within synaptic terminals and chromaffin gland cells is most readily observed in specimens exposed, prior to fixation, to saline solutions containing both tannic acid, and 4-aminopyridine and/or elevated levels of K⁺. These findings show that the pattern of secretory discharge, involving both synaptic and non-synaptic release, which is widespread in invertebrate central nervous systems, is also characteristic of vertebrate, peripheral cholinergic terminals.     

Reversible Inhibition of Acetylcholine Synthesis and Behavioural Effects Caused by 3-Bromopyruvate

3-Bromopyruvate inhibits pyruvate decarboxylase in brain homogenates and causes a 90% drop in acetylcholine tissue content at a concentration of 2 mM. Stereotaxic injection of 3-bromopyruvate into the basal forebrain causes after 7 days a 40% drop of acetylcholine concentration and pyruvate decarboxylase activity in the cortex and hippocampus, and greater decreases at the site of injection. However, values return to normal 18 days after injection. Choline acetyltransferase is partially inhibited only at the site of injection after 7 days. Choline transport and choline concentration are not affected at either 7 or 18 days after injection. Impairments in spontaneous alternation and in retention of passive avoidance were seen only 7 days after the injection. The results suggest that stereotaxic injection of bromopyruvate can induce discrete reversible cholinergic lesions on a time scale useful for behavioural experiments and for comparison with neurodegeneration.     

The regulation and role of neuronal gap junctions during neuronal injury

In the mammalian CNS, excessive release of glutamate and overactivation of glutamate receptors are responsible for the secondary (delayed) neuronal death following neuronal injury, including ischemia, traumatic brain injury (TBI) and epilepsy. The coupling of neurons by gap junctions (electrical synapses) increases during neuronal injury. In a recent study with the use of in vivo and in vitro models of cortical ischemia in mice, we have demonstrated that the ischemic increase in neuronal gap junction coupling is regulated by glutamate via group II metabotropic glutamate receptors (mGluR). Specifically, we found that activation of group II mGluRs increases background levels of neuronal gap junction coupling and expression of connexin 36 (Cx36; neuronal gap junction protein), whereas inactivation of group II mGluRs prevents the ischemia-mediated increases in the coupling and Cx36 expression. Using the analysis of neuronal death, we also established that inactivation of group II mGluRs or genetic elimination of Cx36 both dramatically reduce ischemic neuronal death in vitro and in vivo. Similar results were obtained using in vitro models of TBI and epilepsy. Our study demonstrated that mechanisms for the injury-mediated increase in neuronal gap junction coupling are part of the mechanisms for glutamate-dependent neuronal death.