Failure Of Synaptic Transmission Research Articles

Muscle degeneration following remote nerve injury

Muscle depends upon innervation and contraction to maintain a differentiated state. Denervation can therefore induce muscle atrophy. In grasshoppers, muscle degeneration can also be triggered by the severing of a leg during autotomy. In this case, the muscles that degenerate are neither damaged nor denervated. This phenomenon suggests the existence of transneuronal mechanisms that influence muscle survival. To characterize this autotomy-induced process, we studied the degeneration of a thoracic tergotrochanteral depressor muscle (M#133b,c) subsequent to the shedding of a hindlimb in the grasshoppers Barytettix psolus and Barytettix humphreysii. Both histochemical and electrophysiological methods were used to follow muscle degeneration 1, 3, 5, 10, and 15 days postautotomy. Muscle fibers began to show denervation-like electrophysiological changes (i.e., depolarized resting membrane potentials and postinhibitory rebound) as soon as 3 days postautotomy. By 10 days, significant muscle degeneration was evident and electrophysiological changes were found in all animals tested. Muscle anatomical degeneration was not induced by synaptic transmission failure, because neuromuscular transmission was maintained in most fibers. The rate of muscle degeneration was not constant. Between 1 and 10 days, mean fiber cross-sectional area did not change on the autotomized side, although this is normally a time of muscle growth. However after 10 days, cross-sectional area became drastically reduced and the number of muscle fibers within M#133b,c was decreased. The variability in rate of fiber degeneration was not dependent upon fiber type, since M#133b,c only contains fast-type fibers.

Effects of glucose deprivation in area CA1 of hippocampal slices from adult and juvenile rats

The effects of glucose deprivation were studied in area CA1 of rat hippocampal slices obtained from adult and juvenile rats (postnatal days (PN) 6–8; 13–15; 20–22). Ion-sensitive microelectrodes were employed to monitor baseline and stimulus-induced changes in [Ca 2+] 0, [K +] 0 and field potentials. In slices from juvenile animals, the decline of baseline [Ca 2+] 0 during glucose deprivation was delayed in comparison to adult slices. The minimum in [Ca 2+] 0 was reached in slices from adult rats after 50±8.5 min, in slices from PN 20–22 after 69±9 min, and in slices from PN 13–15 after 111±11 min. In slices from PN 6–8, [Ca 2+] 0 did not decrease significantly even during prolonged exposure of up to 4 h. Alvear stimulation failed to evoke any stimulus-induced responses in field potentials, rises in [K +] 0 and decreases in [Ca 2+] 0 after the minimum in [Ca 2+] 0 was reached in slices from all age groups except for slices from PN 6–8. In the older age groups, afferent fibre stimulation still induced afferent volleys and small decreases in [Ca 2+] 0, which were about 20–30% of those under control conditions, suggesting that presynaptic fibres and endings maintained some of their functional properties even after prolonged glucose deprivation. In contrast, stimulation of the stratum radiatum failed to evoke synaptic responses in slices from PN 6–8, presumably due to a failure in synaptic transmission. These findings confirm that similar to hypoxia during the early postnatal stage, hippocampal neurons are much more resistant to glucose deprivation. The findings also show that during early postnatal development, glucose deprivation may result in a block of synaptic transmission independent of postsynaptic excitability.

Sustained neurotransmitter release: new molecular clues.

Chemical synapses convey impulses at high frequency by exocytosis of synaptic vesicles. To avoid failure of synaptic transmission, rapid replenishment of synaptic vesicles must occur. Recent molecular perturbation studies have confirmed that the recycling of synaptic vesicles involves clathrin-mediated endocytosis. The rate of exocytosis would thus be limited by the capacity of the synaptic clathrin machinery unless vesicles could be drawn from existing pools. The mobilization of vesicles from the pool clustered at the release sites appears to provide a mechanism by which the rate of exocytosis can intermittently exceed the rate of recycling. Perturbation of synapsins causes disruption of vesicle clusters and impairment of synaptic transmission at high but not at low frequencies. Both clathrin-mediated recycling and mobilization of vesicles from the reserve pool are thus important in the replenishment of synaptic vesicles. The efficacy of each mechanism appears to differ between synapses which operate with different patterns of activity.

Altered Na +-channel function as an in vitro model of the ischemic penumbra: action of lubeluzole and other neuroprotective drugs

Veratridine blocks Na +-channel inactivation and causes a persistant Na +-influx. Exposure of hippocampal slices to 10 μM veratridine led to a failure of synaptic transmission, repetitive spreading depression (SD)-like depolarizations of increasing duration, loss of Ca +-homeostasis, a large reduction of membrane potential, spongious edema and metabolic failure. Normalization of the amplitude of the negative DC shift evoked by high K + ACSF 80 min after veratridine exposure was taken as the primary endpoint for neuroprotection. Compounds whose mechanism of action includes Na +-channel modulation were neuroprotective (IC 50-values in μM): tetrodotoxin 0.017, verapamil 1.18, riluzole 1.95, lamotrigine ≥10, and diphenylhydantoin 16.1. Both NMDA (MK-801 and APH) and non-NMDA (NBQX) excitatory amino acid antagonists were inactive, as were NOS-synthesis inhibitors (nitro- l-arginine and l-NAME), Ca 2+-channel blockers (cadmium, nimodipine), and a K +-channel blocker (TEA). Lubeluzole significantly delayed the time before the slices became epileptic, postponed the first SD-like depolarization, allowed the slices to better recover their membrane potential after a larger number of SD-like DC depolarizations, preserved Ca 2+ and energy homeostasis, and prevented the neurotoxic effects of veratridine (IC 50-value 0.54 μM). A concentration of lubeluzole, which was 40× higher than its IC 50-value for neuroprotection against veratridine, had no effect on repetitive Na +-dependent action potentials induced by depolarizing current in normal ACSF. The ability of lubeluzole to prevent the pathological consequences of excessive Na +-influx, without altering normal Na +-channel function may be of benefit in stroke.

Neuronal activity and early neurofibrillary tangles in Alzheimer's disease.

We studied neuronal activity and its relation to the accumulation of neurofibrillary tangles in Alzheimer's disease (AD) neurons by in situ hybridization to cytochrome oxidase subunit III messenger RNA, a marker of mitochondrial energy metabolism. In AD midtemporal cortex, levels of cytochrome oxidase subunit III messenger RNA were decreased by 26% in neurons bearing early-stage neurofibrillary tangles as compared to tangle-free neurons (p < 0.01). However, levels of 12S ribosomal RNA, also encoded by mitochondrial DNA, and of total messenger RNA were decreased only in later stages of tangle development. Comparing tangle-free neurons of 4 AD brains to tangle-free neurons of 3 control brains, levels of cytochrome oxidase subunit III messenger RNA were found to be 25% lower (p < 0.001) in AD tangle-free neurons. Because energy metabolic needs of neurons are mainly determined by synaptic input, the observed decreases in cytochrome oxidase subunit III messenger RNA likely reflect downregulation due to impaired synaptic function in AD. Thus, a failure in synaptic transmission may precede tangle formation. A further decline in neuronal activity is seen as tangle formation progresses. However, these results can also be viewed as showing the viability and continuing activity, albeit at a lower level, of neurons in the early stages of neurofibrillary pathology.

Effects of cerebral ischemia on N-methyl-D-aspartate and dihydropyridine-sensitive calcium currents. An electrophysiological study in the rat hippocampus in situ.

During cerebral ischemia, both promoting and limiting factors are present for activation of the N-methyl-D-aspartate (NMDA) receptor ion channel and the dihydropyridine (DHP)-sensitive Ca2+ channels. We investigated the activity of these channels during ischemia and reperfusion in the rat hippocampus in situ. Reversible ischemia was induced by bilateral carotid artery ligation. NMDA and BAY K8644 were applied by iontophoresis or pneumatic ejection, and extracellular field potential and resistance changes were recorded from the CA1 region of the rat hippocampus. Resting membrane potentials of the CA1 neurons were also recorded. DC potential shifts produced by NMDA and BAY K8644 were reduced when ischemia depressed the evoked activity more than 50%. They disappeared on total failure of synaptic transmission and recovered during reperfusion. When the evoked activity was depressed less than 50%, DC shifts were greater than their preischemic values; however, BAY K8644-induced potentiation did not reach statistical significance. CA1 neurons were depolarized during ischemia. These data suggest that ischemia severe enough to cause transmission failure inactivates NMDA and DHP-sensitive Ca2+ currents. During less intense ischemia and reperfusion, NMDA and DHP-sensitive Ca2+ channels are functional, and their overactivation may lead to neurotoxicity.

Distinct Roles for N-Ethylmaleimide-sensitive Fusion Protein (NSF) Suggested by the Identification of a Second Drosophila NSF Homolog

The N-ethylmaleimide-sensitive fusion protein (NSF) is a cytoplasmic protein implicated in the fusion of intracellular transport vesicles with their target membranes. NSF is thought to function in the fusion of essentially all types of vesicles, including endoplasmic reticulum, Golgi, and endocytic vesicles, as well as secretory vesicles undergoing regulated fusion (for review see Rothman, J.E. (1994) Nature 372, 55-63). However, little experimental evidence exists to address the possibility that organisms might have multiple NSF proteins serving distinct functions in the same or different cells. We previously cloned a neurally expressed Drosophila homolog, dNSF-1 (Ordway, R.W., Pallanck, L., and Ganetzky, B. (1994) Proc. Natl. Acad. Sci. U.S.A. 91, 5715-5719), and have subsequently identified mutations in this gene that confer an apparent failure of synaptic transmission at elevated temperature (Pallanck, L., Ordway, R.W., and Ganetzky, B. (1995) Nature, 376, 25; Siddiqi, O., and Benzer, S. (1976) Proc. Natl. Acad. Sci. U.S.A. 73, 3253-3257). Here we report that 1) Drosophila contains a second NSF homolog, termed dNSF-2, that exhibits 84% amino acid identity to dNSF-1, 2) dNSF-1 and dNSF-2 display overlapping but different temporal expression, and 3) multiple transcripts are derived from the dNSF-2 gene. These findings raise the possibility that different NSF gene products serve distinct or overlapping functions with the organism.

Intracellular calcium levels and calcium fluxes in the CA1 region of the rat hippocampal slice during in vitro ischemia: relationship to electrophysiological cell damage

Five minutes of oxygen and glucose deprivation (termed "in vitro ischemia") causes long-term synaptic transmission failure (LTF) in the CA1 region of the rat hippocampal slice. Dependence of LTF on cell calcium was tested by generating graded reductions in cell Ca. There was a strong correlation between the average level of exchangeable cell Ca in CA1 during ischemia, and the extent of LTF. In standard buffer, exchangeable cell Ca in CA1 increased by 35% after 3 min of ischemia and remained elevated for the entire 5 min of ischemia. Unidirectional Ca influx increased by 35% during the first 2.5 min of ischemia and remained at that level for the next 2.5 min. There were no changes in unidirectional Ca efflux during this period. Thus, the accumulation results from increased influx of Ca. Ca influx during the first 2.5 min of ischemia depended entirely on NMDA channels; it was completely blocked by the noncompetitive NMDA receptor antagonist MK-801. However MK-801 had no effect during the second 2.5 min. This inactivation of NMDA-mediated influx during ischemia appears to result from dephosphorylation. Okadaic acid increased Ca influx during the second 2.5 min of ischemia and this increase was blocked by MK-801. The ischemia-induced Ca influx during the second 2.5 min of ischemia was attenuated 25% by nifedipine (50 microM) and an additional 35% by the Na/Ca exchange inhibitor benzamil (100 microM). The AMPA/kainate antagonist DNQX had no effect on the Ca influx. Antagonists were used to relate Ca influx to LTF. Blockade of enhanced Ca entry during ischemia in standard buffer (2.4 mM Ca) had no effect on LTF, consistent with total cell Ca prior to ischemia being adequate to cause complete LTF. However, MK-801 strongly protected against LTF when the buffer contained 1.2 mM Ca, a more physiological level. MK-801 combined with DNQX prevented transmission damage in standard buffer. Thus, AMPA/kainate receptor activation contributes to ischemic damage, although not by enhancing Ca entry.

Effects of prolonged elevation of potassium on hippocampus of anesthetized rats

We tested the after-effects of prolonged depolarization on neurons in situ in the mammalian brain and examined the site of blockade responsible for failure of synaptic transmission. The CA1 region of the hippocampus of anesthetized rats was exposed to solutions containing elevated concentration of K + (100–125 mmol/1), administered either by microdialysis in intact brain or by irrigation of the exposed surface of the hippocampus. Recovery was observed for 5–61/2h. When K + was administered by microdialysis, evoked potentials were recorded from points near (up to 0.2 mm) and far (0.7–1.0 mm) from the dialysis probe. High K + dialysis induced recurrent waves of spreading depression and, in about half of the preparations, a prolonged unstable depressed state. In the intervals between SD waves orthodromic but not antidromic population spikes remained severely depressed at the ‘far’ recording site. Following high K + dialysis orthodromic population spikes recovered in a triphasic cycle: partial recovery with hyper-transmission was followed bysecondary depression and finally by slow partial or complete recovery. Final recovery was less complete in cases that have experienced prolonged spreading depression. Current source density analysis revealed that during secondary depression transmission was blocked due to failure of dendritic action potentials. When the exposed hippocampus was irrigated with high K + solution ortho- and antidromic evoked potentials recovered completely following high K + exposure of less than 30 min, incompletely after 45 min and failed entirely after 60 min. We conclude that prolonged steady depolarization of hippocampal CA1 pyramidal neurons causes lasting loss of function. Dendritic function is especially prone to depolarization-induced injury. CA1 neurons are less vulnerable in situ than they are in vitro.

Adenosine release is a major cause of failure of synaptic transmission during hypoglycaemia in rat hippocampal slices

Glucose-free medium (aglycaemia) caused a complete failure of CA1 population spikes (after 14.5 ± 0.8 min) and field EPSPs (after 18.1 ± 0.5 min). In the presence of the selective adenosine Al antagonist, 8-( p-sulfophenyl)theophylline (10 μM), population spikes and EPSPs were decreased by only 13.8 ± 11.9% and 32.4 ± 11.6% at the end of 17.0 ± 3.0 min and 19.8 ± 1.7 min of aglycaemia, respectively. A similar effect was produced by caffeine (0.2 mM). The ATP-sensitive K + channel blockers tolbutamide (1 mM) and glibenclamide (10 μM) had no significant effect on aglycaemic suppression of synaptic transmission. These observations indicate that endogenous adenosine, but not ATP-sensitive K + conductance, plays a major role in hypoglycaemia failure of transmission.

Suppression of presynaptic calcium currents by hypoxia in hippocampal tissue slices

We tested the hypothesis that suppression of inward calcium current in presynaptic terminals is the cause of failure of synaptic transmission early during cerebral hypoxia. Postsynaptic responses in CA1 zone of hippocampal tissue slices were blocked either by the combined administration of 6,7-dinitroquinoxaline-2,3-dione (DNQX) and3-((±)-2-carboxypiperazine-4-yl)-propyl-1-phosphonic acid (CPP) or by lowering extracellular calcium concentration ([Ca 2+] 0). Repetitive orthodromic activation of central neurons caused transient decrease of [Ca 2+] 0 (measured by ion selective microelectrodes) in neuropil, attributable to influx of Ca 2+ in presynaptic terminals. Presynaptic [Ca 2+] 0 responses were rapidly and reversibly suppressed when oxygen was withdrawn from hippocampal tissue slices. The ‘resting’ baseline level of [Ca 2+] 0 declined at first gradually, then precipitously as in spreading depression (SD). Presynaptic volleys during high frequency train stimulation were also depressed somewhat before SD began. We conclude that (1) presynaptic Ca 2+ currents fail during hypoxia, perhaps because ‘resting’ intracellular free Ca 2+ activity is increased and, in part, also because of partial failure of presynaptic impulse conduction; (2) the influx of Ca 2+ into brain cells in hypoxic spreading depression is not mediated by glutamate/aspartate dependent channels.

Hypoxic failure of synaptic transmission in the isolated spinal cord and the effects of divalent cations

Responses evoked by stimulation of a dorsal root were recorded from ventral and dorsal roots of isolated spinal cords of infant mice. Interstitial potassium, [K +] o, and extracellular DC voltage were recorded from dorsal gray matter in some experiments. When oxygen was withdrawn, synaptically transmitted discharges (dorsal horn response, DHR, and monosynaptic ventral root reflex, VRR) began to be depressed within a minute, and were depressed to less than 30% of control amplitude in 10–15 min. Responses recovered fully if oxygen was readmitted within 45 min, but no recovery was seen after 90 min of hypoxia. The degree of the depression of VRR was as expected from the depression of the electronically conducted excitatory postsynaptic potential (VRepsp). Responses failed much more rapidly in spinal cords of 15–16-day-old mice, than of 9–14-day-olds. When the spinal cord was bathed in elevated [Ca 2+] o or in reduced [Mg 2+] o, synaptic transmission was consistently maintained for a longer period of hypoxia than in bathing fluid of normal cation content. In a sizeable minority of the trials during hypoxia an abrupt increase of [K +] o occurred, accompanied by a sudden negative shift of extracellular potential, closely resembling spreading depression (SD) of forebrain structures. Delayed post-hypoxic spontaneous activity was seen in many spinal cords. The results are compatible with the hypothesis that hypoxic failure of synaptic transmission is due, in part or whole, to blockade of inward Ca 2+-current in presynaptic terminals. Cells in spinal gray matter can no longer be regarded as ‘immune’ to SD-like depolarization, but the limited conditions under which SD can occur are not yet clear.

Effects of chronic in vivo replacement of choline with a false cholinergic precursor.

Chronic administration to rats of a diet in which all choline is replaced by NADe, an unnatural choline analog, results in a classical hypocholinergic syndrome characterized by progressive loss of learning and memory, hyperkinesis, hyperreactivity and hyperalgesia. Discontinuation of the artificial diet results in rapid elimination of NADe from both free and phospholipid-bound pools in all tissues studied, but the behavioral effects recede more slowly and incompletely. These results are consistent with a model in which choline and NADe compete in both acetylcholine and phospholipid synthesis, resulting in selective vulnerability of cholinergic neurons. Histological studies are in progress to determine whether microanatomical changes are also consistent with this model.

ATP depletion in slow-twitch red muscle of rat.

Rat slow-twitch muscle, in contrast to fast-twitch muscle, maintains its ATP content near normal during intense stimulation conditions that produce rapid fatigue. An extensive depletion of adenine nucleotide content by the deamination of AMP to IMP + NH3, typical of fast-twitch muscle, does not occur. We evaluated whether this response of slow-twitch muscle could be simply due to failure of synaptic transmission or related to cellular conditions influencing enzyme activity. Stimulation of soleus muscles in situ via the nerve or directly in the presence of curare at 120 tetani/min for 3 min resulted in extensive fatigue but normal ATP contents. Thus the lack of ATP depletion must be related to cellular events distal to neuromuscular transmission. Even nerve and direct muscle stimulation (with curare) during ischemia did not cause a large depletion of ATP or a large elevation of lactate content (12.0 +/- 0.7 mumol/g), even though the decline in tension was essentially complete. However, if the same tension decline during ischemia was prolonged by stimulating for 10 min at 12 tetani/min a large decrease in ATP (2.24 +/- 0.09 mumol/g) and increase in IMP (2.47 +/- 0.16 mumol/g) and lactate (30.4 +/- 2.0 mumol/g) content occurred. Thus adenine nucleotide deamination to IMP can occur in slow-twitch muscle during specific contraction conditions. The cellular events leading to the activation of AMP deaminase require an intense contraction condition and may be related to acidosis caused by a high lactate content.

Extracellular potassium ion activity and electrophysiology in the hippocampal slice: paradoxical recovery of synaptic transmission during anoxia

The relationship between extracellular potassium ion activity and neuronal excitability during anoxia was investigated in hippocampal slices in vitro. Extracellular field potentials and K + activity were measured with double-barreled ion-selective microelectrodes placed either in the stratum pyramidale or stratum radiatum of field CA 1. Orthodromic spike ctivity of CA 1 pyramidal cells and field excitatory postsynaptic potentials (f-EPSPs) failed rapidly after anoxia with little change in potassium ion activity and without failure of the Schaffer collateral prevolley or antidromic responses of pyramidal cells. As [K +] 0 approached 8–10 mM, f-EPSPs and orthodromic spike activity recovered spontaneously. Continued anoxia resulted in massive release of K + into extracellular space and complete electrical silence. Presynaptic activity and antidromically elicited spike activity recovered promptly upon reoxygenation after anoxia, but synaptic transmission remained blocked for many minutes. Spontaneous recovery of f-EPSPs and spike activity suggests that a simple mechanism involving depolarization of hyperpolarization of neuronal elements cannot account for failure of synaptic transmission observed during anoxia. However, continued elevation of [K +] 0 and the associated loss of pre- and postsynaptic excitability with more prolonged anoxia indicated that depolarization was responsible for the eventual electrical silence as anoxia progressed.

Noradrenalin-inducible cyclic-AMP accumulation in rat cerebral cortex: changes during complete global ischemia.

Neurologic dysfunction after cerebral ischemic insults may be due not only to neuronal death, but also to a possibly reversible failure in synaptic transmission. Because noradrenaline (NA)-inducible cyclic-AMP (cAMP) accumulation in brain may reflect the integrity of synaptic transmission mechanisms and brain viability, we studied its changes in cerebral cortex after various durations of decapitation ischemia. Unanesthetized rats were decapitated and the brains were kept at 37 degrees C for times ranging from 0 to 60 min. Cerebral cortical slices were incubated in vitro and NA (11.2 microM)-induced cAMP accumulation was evaluated over 10 min. At 0 min of ischemia, NA-induced cAMP accumulation was 56 pmol/mg protein/10 min. Between 0 and 20 min of ischemia, a linear eightfold increase, to 435 +/- 49 pmol/mg protein/10 min, occurred in NA-induced cAMP accumulation, with no further increase after longer durations of ischemia. The mechanisms modulating the increase in cortical NA-inducible cAMP accumulation with a maximum response after 20 min of ischemia remain to be defined.

Reduced ATP concentration as a basis for synaptic transmission failure during hypoxia in the in vitro guinea-pig hippocampus.

1. Experiments were performed to determine whether a decrease in tissue ATP contributes to the rapid failure of cerebral synaptic transmission during hypoxia. Transmission between the perforant path and the dentate granule cells in the in vitro hippocampus was studied.2. Hippocampal slice ATP is decreased by approximately 15% at the time that the evoked response begins to diminish in standard Krebs bicarbonate buffer. This is about 2 min after the onset of hypoxia.3. When transmission failure is accelerated by increasing extracellular K(+) from 4.4 to 13.4 mM, the evoked response begins to decay about 30 sec after exposure to hypoxia. There is no decrease in hippocampal slice ATP at this time.4. However, ATP in the molecular layer (the synaptic region of the tissue) is decreased by approximately 15% at the time the evoked response begins to decay in the slices exposed to elevated K(+) concentration.5. Exposing the hippocampal slice to 25 mM-creatine for 3 hr elevates molecular layer phosphocreatine fourfold. Synaptic transmission during hypoxia survives three times as long as it does in the absence of creatine.6. In the creatine fortified medium, molecular layer ATP no longer declines within 30 sec of hypoxia. However the molecular layer ATP does decline within 90 sec of hypoxia, the time at which the evoked response begins to decay in this creatine-fortified buffer.7. The results establish that ATP in the region of the active synapses is lowered when the first signs of electrophysiological failure appear during hypoxia. They also show that maintaining ATP for longer than normal during hypoxia is associated with a prolonged maintenance of the evoked response. They thus suggest that a decline in ATP is one factor causing hypoxic block of synaptic transmission.8. It is further suggested that the very rapid failure of the electroencephalogram during anoxia may also result from a decline in ATP.

The Effects of Cooling On an Identified Reflex Pathway in the Cockroach (Periplaneta Americana), in Relation to Chill-Coma

ABSTRACT The effects of cooling were studied in relation to (a) the impairment of locomotion (i.e. chill-coma) and (b) the functioning of various components of the monosynaptic trochanteral hair plate reflex in the cockroach, Peri-planeta americana. The mean temperature for onset of chill-coma was 10·5 °C. At this temperature animals were unable to right themselves and visible tremors of the legs and body occurred. Extracellular recordings from metathoracic nerve 5 indicated that cooling from 24 to 15 °C caused a decrease in the background spiking rate of motor neurone Ds. However, cooling from 12 to 9 °C caused a marked increase in the spiking frequency of both Ds and other unidentified neurones. This increase in spiking activity is the probable basis of leg and body tremors that occur during chill-coma. Monosynaptic coupling between hair plate afferent spikes and Ds spikes (following electrical stimulation of the hair plate) was markedly affected by cooling. At 25 °C, hair plate afferent and Ds spikes were always tightly coupled to one another, whereas at 10 °C coupling was very weak. Several lines of evidence suggest that this loss of coupling was the result of failure of central synaptic transmission. The reduced effectiveness of central synaptic transmission during cooling may be a critical factor in the impairment of locomotor ability associated with chill-coma.

Cerebral synaptic transmission during anoxia is protected by creatine.

Synaptic transmission in cerebral tissue fails very rapidly in the absence of oxygen; the metabolic basis for this is not known. We report here that the transmission failure in the guinea pig hippocampal slice can be delayed threefold by exposing the tissue to extracellular creatine (Cr) for 3 h. The improved survival is associated with an increase of tissue phosphocreatine (PCr) concentration. These data argue that the metabolic basis for synaptic transmission failure is a fall in tissue ATP concentrations. They also indicate a way to protect brain tissue against anoxic damage.

Blocking action of intracellularly injected neuraminidase on central synapses in vivo.

The effect of neuraminidase on synaptic transmission was studied at cholinergic and noncholinergic contacts in the buccal and cerebral ganglion of Ap]lysia. The amplitudes of monosynaptic unitary postsynaptic potentials generated by intracellular stimulation of identified presynaptic neurones were measured as indication for the efficacy of synaptic transmission. Neuraminidase was either intrasomatically injected into a presynaptic neurone, or the whole ganglion was incubated with the enzyme. Intrasomatic injection of the enzyme resulted in complete failure of synaptic transmission. This effect occurred independently of the transmitter used. The synaptic failure was presynaptic in origin. The biophysical characteristics of an injected neurone, particularly the amplitude and propagation of its action potential, did not appear to be affected by neuraminidase. Synaptic transmission and biophysical membrane properties were unaffected by extracellular neuraminidase. We conclude that the synaptic blockade is due to the enzyme's action inside the presynaptic nerve ending. It seems most likely that neuraminidase cleaves sialic-acid-containing-compounds associated with the nerve terminal surface membrane, probably thus causing failure of transmitter release.