Possible role of prostaglandins in post-tetanic potentiation at the nerve-muscle junction in the longitudinal muscle strip of guinea-pig ileum

The effect of tetanic stimulation on the twitch responses of the longitudinal muscle-myenteric plexus preparation of the guinea-pig ileum to electrical stimulation was investigated in the presence of naloxone. Under this condition, or after the addition of PGE2, twitch contractions were maximal and no potentiation of twitches following tetanus was observed. In the presence of indomethacin (1 mumol litre-1) twitches were diminished and post-tetanic potentiation (PTP) was manifested. PTP was seen with indomethacin concentrations of 1 to 20 mumol litre-1 or after simultaneous addition of diphloretin phosphate (16 mumol litre-1). Thus it seems unlikely that the effect of prostaglandins released during tetanic stimulation would be of key importance for the manifestation of PTP. Rather it is thought that a decrease in the release of acetylcholine from motor nerve terminals, and consequently smaller twitches in the presence of indomethacin, offer favourable conditions for PTP.     

The influence of ouabain on twitch contractions in the presence of veratridine

1. Direct stimulation evoked twitches in mouse diaphragm muscles in presence of 10 microM D-tubocurarine in vitro. Effects of ouabain and their dependence on K+ were examined on the twitch responses and action potentials in the presence and absence of twitch potentiators. 2. Ouabain inhibited twitch contractions only in the presence of veratridine, aconitine and monensin while it had no inhibitory effect on control twitches. The interactions between ouabain and these twitch potentiators depended on the presence of external K+, except in the case of monensin. 3. Removal of Ca2+ from a bathing solution accelerated the potentiating effect of veratridine and the antagonizing effect of ouabain. 4. Caffeine further potentiated the twitches which had been attenuated by ouabain combined with veratridine. 5. Ouabain combined with veratridine consistently decreased resting membrane potentials, action potentials and overshoot potentials and prolonged time to peak of and duration of the muscle action potentials. 6. Tetraethylammonium, 4-aminopyridine, and caffeine produced twitch potentiation which was insensitive to ouabain or the removal of K+. 7. These results suggest that twitch contractions in the presence of activators of sodium channels link with activation of Na+-K+-ATPase. Accumulation of Na+ inside the muscle fibres may uncouple the excitation-contraction system. 8. This uncoupling may not include the caffeine-sensitive process that controls the release of Ca2+ from the sarcoplasmic reticulum. Na+ accumulation may decrease transmembraneous gradient of this ions, thereby causing a reduction in excitation coupled with twitch contraction.     

A STUDY OF STIMULATION-EVOKED ACTIVATION OF α2-ADRENOCEPTORS IN THE RAT ISOLATED VAS DEFERENS

Experiments were performed with prostatic and epididymal segments of rat vas deferens to determine whether alpha 2-adrenoceptors in this organ were activated during field stimulation of sympathetic terminals with short trains of pulses applied at low frequencies. In prostatic segments the magnitude of twitches evoked by trains of ten field pulses at 1 or 2 Hz declined after 2-3 s of stimulation. In contrast, facilitation of twitches and fusion of contractions occurred when similar stimulation was applied to epididymal segments. In prostatic segments from rats treated with reserpine (2.5 mg/kg i.p.) 24 h previously, there was no decline in the magnitude of twitches produced by successive impulses in a train of stimulating pulses. In epididymal segments from reserpine-treated rats facilitation of twitches in response to successive impulses in each train still occurred. In prostatic segments cocaine (5 and 10 mumol/l) enhanced twitch fade with stimulation at 1 and 2 Hz without altering the time for onset of this effect. In epididymal segments cocaine led to enhancement and prolongation of contractile responses. In prostatic segments yohimbine (0.01-0.06 mumol/l) reduced or reversed the effect of cocaine in enhancing twitch fade. In preparations where the reversal of the effect of cocaine by yohimbine was incomplete, subsequent addition of phentolamine (1 mumol/l) produced complete reversal. In epididymal segments yohimbine (0.01 mumol/l) produced a further enhancement in the twitch responses to stimulation at 1 and 2 Hz. Subsequent addition of phentolamine (1 mumol/l) reversed the facilitatory effects of cocaine and yohimbine. Propranolol (10 mumol/l) was without effect upon responses to stimulation in either segment of rat vasa deferentia. These experiments indicate that noradrenaline, released by short trains of impulses applied at low frequency to hypogastric nerve terminals activates prejunctional alpha 2-adrenoceptors in the prostatic segment of the vas deferens. In the epididymal portion the effects arising from activation of prejunctional alpha 2-adrenoceptors are outweighed by the consequences of activation of extrajunctional alpha 1-adrenoceptors located on longitudinally arranged muscle.     

Endogenous adenosine prevents post-tetanic release facilitation mediated by alpha3beta2 nicotinic autoreceptors

We investigated the modulatory role of endogenous adenosine on tetanic-induced (50 Hz for 5 s) nicotinic facilitation of [3H]acetylcholine release (5 Hz for 50 s) from rat motoneurons. Adenosine deaminase (0.5 U/ml) and the adenosine A(2A) receptor antagonist, 3,7-dimethyl-1-propargyl xanthine (DMPX, 30 microM), facilitated post-tetanic [3H]acetylcholine release. Release inhibition caused by tubocurarine (1 microM), dihydro-beta-erythroidine (1 microM) and alpha-conotoxin MII (0.1 microM) was attenuated after tetanic preconditioning. Nicotinic inhibitory action was fully restored after adenosine A(2A) receptor block by DMPX or adenosine deaminase. DMPX (10 microM) caused a leftward shift of the inhibitory dose-response curves for d-tubocurarine (0.1-1 microM), dihydro-beta-erythroidine (0.03-10 microM) and alpha-conotoxin MII (1-300 nM) on post-tetanic twitch amplitude. In contrast, the post-tetanic twitch depression caused by alpha-bungarotoxin (3-100 nM, which had no effect on transmitter release) was attenuated by DMPX (10 microM). It is concluded that activation of adenosine A(2A) receptors by endogenously generated adenosine prevents the post-tetanic release facilitation mediated by nicotinic alpha3beta2 autoreceptors.     

The effects of tetraethylammonium during twitch and tetanic stimulation of the phrenic nerve diaphragm preparation in the rat

Tetraethylammonium (TEA) (2.6 x 10(-3) M) potentiated the twitches of the indirectly- or directly-stimulated phrenic nerve diaphragm of the rat at 37 degrees C by prolonging the action potential of the sarcolemma, due to an inhibition of the repolarizing K+ current. With indirect stimulation, TEA caused a use-dependent inhibition of tetanic contractions, induced every 10 min by 10 sec of 50 Hz stimulation, and a post-tetanic depression of the twitches was observed after about 40 min. Recording of the electromyogram (EMG) and compound action potentials of the phrenic nerve, localized the two inhibitory effects to the neuromuscular junction. They were caused by different mechanisms of action. Choline (3.6 x 10(-4) M) antagonized the depression of the twitch but not the use-dependent inhibition. Lowering the temperature to 20 degrees C reduced the depression of the twitch, whereas the use-dependent inhibition was enhanced. The release of transmitter was probably normal during tetanic stimulation; a post-synaptic desensitization of acetylcholine (ACh) receptors caused the inhibition. Microelectrode recordings of endplate potentials supported this conclusion. The depression of the twitch was due to a presynaptic depletion of transmitter. This was confirmed by inducing an additional depletion and depression of the twitch with N-ethyl-maleimide (2.5 x 10(-5) M). Since the depression of the twitch was antagonized by choline, the depletion was probably due to an inhibited uptake of choline into the nerve terminals.     

Effects of atropine and glycopyrrolate on neuromuscular transmission in the rat phrenic nerve-diaphragm preparation

1. The effects of atropine and glycopyrrolate on neuromuscular transmission and on muscle contraction, were studied, in the rat diaphragm preparation, by analyzing their effects on the indirectly (and directly)-elicited twitch (0.2 Hz), tetanic (50 Hz for 20 sec duration), post-tetanic twitch responses (at 5 sec after the tetanus), and on the phenomenon of post-tetanic twitch potentiation (PTP), which is thought to be of a presynaptic origin, i.e. due to increased transmitter release. 2. Atropine (0.001-10 microM) increased the indirectly-elicited twitch tension by 22 +/- 2.1% (control 0.9 +/- 0.1 g, P less than 0.02), the tetanus by 15 +/- 1.1% (control 3.9 +/- 0.7 g, P less than 0.05), the post-tetanic twitch response by 33 +/- 3.1% (control 1.2 +/- 0.1 g, P less than 0.01) and the PTP value by 36 +/- 1.9% (control 33 +/- 2.3%, P less than 0.01, means +/- SEM = 6). 3. Atropine (0.001-10 microM) had little effect on the directly-elicited twitch tension, but in high concentrations (e.g. 20 microM), it blocked the twitch tension. 4. In contrast, glycopyrrolate (0.1-100 microM) had little effect on the twitch tension (direct or indirect), but it significantly reduced the tetanus (by 38 +/- 3.5%, P less than 0.01), the post-tetanic twitch response (by 17 +/- 1.2%, P less than 0.05) and the PTP values (by 24 +/- 3.1% P less than 0.02). 5. In the presence of hemicholinium (1.3 microM) the responses to atropine and glycopyrrolate were altered (decreased), indicating a possible action on presynaptic mechanism of transmission. 6. It is concluded that atropine and glycopyrrolate produce different (opposite) effects at the rat neuromuscular junction, atropine enhances whereas glycopyrrolate depresses neuromuscular transmission. The effects of these two antimuscarinic drugs may be exerted at the presynaptic nerve terminals, i.e. on presynaptic muscarinic receptors, which are involved in the feedback mechanism of transmitter release.     

Antagonistic action of uranyl nitrate on presynaptic neurotoxins from snake venoms

Uranyl ion (UO2+2) antagonized the neuromuscular blocking action and phospholipase A2 activity of neurotoxins which act presynaptically [beta-bungarotoxin (beta-BuTX) and crotoxin] but did not affect the action of alpha-bungarotoxin and tetrodotoxin. On the basis of the kinetic analysis of the UO2+2 and strontium ion (Sr2+) antagonism of muscle paralysis induced by beta-bungarotoxin, it was found that they inhibited both the binding of the toxin and the steps following binding that brought about the neuromuscular blocking action of beta-bungarotoxin. Uranyl ion was about 50 times more potent than Sr2+ in antagonizing beta-bungarotoxin. High Ca2+ (10 mM) abolished but low Ca2+ (0.25-1.25 mM) medium enhanced the antagonizing action of UO2+2 and Sr2+. In low Ca2+ medium, UO2+2 markedly potentiated the amplitude of the twitch, subsequent addition of beta-bungarotoxin produced three phases of effects on the twitches, e.g. an initial depression, followed by the second facilitation and finally a rapid depression of twitches; however, approx. 70 min after beta-bungarotoxin the small twitches reached a steady state which persisted for more than 350 min. Therefore, it is evident that UO2+2 is the most potent antagonist of beta-bungarotoxin so far tested.     

Neuromuscular blocking profile of the vecuronium analogue, Org-9487, in the rat isolated hemidiaphragm preparation.

1. The neuromuscular effects of the short-acting aminosteroid muscle relaxant Org-9487 have been studied in the in vitro rat phrenic nerve/hemidiaphragm preparation by use of twitch tension and electrophysiological recording techniques. 2. Org-9487 (5-100 microM) produced a concentration-dependent decrease in the amplitude of twitches (0.1 Hz) and tetanic contractions (50 Hz) evoked by motor nerve stimulation. The compound produced fade of force during both 50 Hz stimulation and train-of-four stimulation at 2 Hz, indicating a prejunctional component of action. 3. Anticholinesterases only partially reversed the effect of Org-9487 on twitch responses. This was possibly because, at the concentrations required to block twitches in the rat, Org-9487 itself was found to possess significant anticholinesterase activity. 4. Org-9487 (3 microM) increased the rundown of endplate current amplitudes during a 2 s train of 50 Hz nerve stimulation. This was because Org-9487 increased the quantal content of the first endplate current in the train without affecting acetylcholine release towards the latter part of the train. 5. Org-9487 (10 microM) produced a voltage-dependent decrease in the time constant of decay of endplate currents at 32 degrees C and 0.5 Hz, indicative of a block of endplate ion channels. The blocking rate constant increased with membrane hyperpolarization.     

Dependence on temperature of the effect of dinitrophenol on the release of transmitter quanta at neuromuscular junctions in the mouse diaphragm.

1. The frequencies (F, s-1) of miniature endplate potentials and the quantal content (m) of endplate potentials were measured intracellularly and simultaneously at mouse diaphragm endplates in a bathing solution that contained 0.6 mM Ca2+ ions and 5 mM Mg2+ ions. 2. Twin pulses at 4 ms intervals gave the quantal contents of the first (m1) and second (m2) responses. The ratio of m2/m1 was taken as an indicator of the temporal facilitation of the release of transmitter. 3. Dinitrophenol (DNP, 10 microM) increased the values of F and m at 36 degrees C. This effect did not depend on extracellular Ca2+ ions. 4. The potentiating effect of DNP disappeared at 24 degrees C but the value of m2/m1 remained constant. 5. These results suggest that the effect of DNP is modifiable by temperature which can affect systems that control the intracellular metabolism of Ca2+ ions.     

The influence of ouabain on twitch potentiation by anticholinesterases in the phrenic nerve-diaphragm muscles of mice.

1. (+)-Tubocurarine, hexamethonium, atropine, ouabain, and removal of potassium from the bathing medium were examined for their effects on indirectly evoked twitches (IT) of mouse phrenic nerve-diaphragm muscles in the presence or absence of neostigmine. 2. Neostigmine increased the amplitude of IT. The twitch potentiation was reduced by (+)-tubocurarine at low concentrations that had no inhibitory effect on normal IT. Hexamethonium (10-100 microM), but not atropine (0.1-1 microM), partially inhibited the twitch potentiation. Neither hexamethonium nor atropine had an inhibitory effect on IT in the absence of neostigmine. 3. Ouabain (5 microM) abolished the twitch potentiation by neostigmine while having no inhibitory effect on directly evoked twitches in the presence of neostigmine and (+)-tubocurarine together. 4. The potentiating effect of neostigmine was less in a potassium-free bathing solution. The inhibitory effect of ouabain disappeared in this solution. 5. Reinclusion of KCl at 2.5 mM restored both the potentiating effect of neostigmine and the antagonistic effect of ouabain. This reinclusion did not potentiate IT in the absence of neostigmine. 6. An interaction resembling that between ouabain and neostigmine was obtained between ouabain and physostigmine or paraoxon. 7. Both endplate potentials (e.p.ps) and miniature e.p.ps increased in terms of their amplitude and duration in the presence of neostigmine. Ouabain did not reduce the enhanced endplate responses. 8. These results indicate that the potentiation of IT by anticholinesterases may occur via nicotinic receptors which are sensitive to both (+)-tubocurarine and hexamethonium, and that the interaction between anticholinesterases and ouabain depends on the presence of K+. It appears that the mechanisms of twitch potentiation are dependent on the ionic gradients maintained by Na+-K+-ATPase.     

TETANIC FADE DURING PARTIAL TRANSMISSION FAILURE PRODUCED BY NON-DEPOLARIZING NEUROMUSCULAR BLOCKING DRUGS IN THE CAT

1. A comparison has been made of the effects of three acetylcholine antagonists—hexamethonium, tubocurarine and pancuronium—on maximal tetani of limb muscles of cats under chloralose anaesthesia. In most experiments, the indirectly stimulated soleus muscle was studied, but observations were also made on the tibialis anterior and flexor digitorum longus muscles. 2. When neuromuscular block was produced by intra-arterial injections of the acetylcholine antagonists, tetanic tension, though depressed in amplitude, did not wane and there was little or no post-tetanic relief of the block as judged by the amplitude of subsequent twitches. On the other hand, during similar degrees of block produced by intravenous injections, tetanic tension rapidly waned, and, after the tetanus, transmission was temporarily facilitated, as evidenced by an increase in the amplitude of post-tetanic twitches. 3. Intravenously injected hexamethonium caused complete waning of tetanic tension in doses too small to depress twitch amplitude and which caused only a small depression of peak tetanic tension. In contrast, pancuronium caused only partial tetanic fade even in doses that produced pronounced depressions of twitch and tetanic tensions. The effects of tubocurarine fell between these extremes. 4. The results suggest that depression of peak tension and tetanic fade are independent effects of acetylcholine antagonists. It is postulated that the former is a consequence of block of post-junctional cholinoceptors, whereas the latter arises from an action at pre-junctional cholinoceptors. 5. The results obtained, together with those of other workers, led to the suggestion that transmitter acetylcholine, in addition to evoking the endplate potential, acts on the nonmyelinated nerve terminals in a positive feed-back mechanism that mobilizes transmitter to keep pace with release during high frequencies of stimulation.     

Mechanisms underlying the vecuronium-induced tetanic fade in the isolated rat muscle

The cellular mechanisms underlying the effects of vecuronium on the tetanic contraction were studied in vitro with a combination of myographic and electrophysiologic techniques. We used the isolated sciatic nerve extensor digitorum longus muscle preparation of the rat. Indirect twitches were evoked at 0.1 Hz pulses and tetani at 50 Hz pulses. Trains of end-plate potentials were generated at 50 Hz. The electrophysiological variables used in the analysis of the end-plate potentials were: amplitude, tetanic run-down, quantal size and quantal content. The myographic study demonstrated that vecuronium at 0.4 microM caused tetanic fade, but left the twitch unaffected. Regarding electrophysiology, vecuronium (0.4 microM) decreased the amplitude of end-plate potentials and increased their tetanic run-down. These changes were due to significant reductions in both the quantal content of the end-plate potentials and the quantal size. It is concluded that vecuronium has both pre- and postsynaptic effects at the neuromuscular junction, and that it induces fade of the tetanic contraction via a summation of these effects.     

The effect of ions and second messengers on long-term potentiation of chemical transmission in avian ciliary ganglia.

1. The effects of tetanic stimulation of the oculomotor nerve on transmission through the avian ciliary ganglion have been determined by use of the amplitude of the compound action potential recorded in the ciliary nerve, in the presence of hexamethonium (300 microM), as a measure of synaptic efficacy. 2. Tetanic stimulation for 20 s at 30 Hz potentiated the chemical phase of the compound action potential by at least 100% of its control level. This potentiation, reflecting an increase in synaptic efficacy, decayed over two distinct time courses: firstly, a rapid decay with a time constant in the order of minutes, and secondly, a slower decay, representing a smaller potentiation, with a time constant in the order of an hour. The large increase in synaptic efficacy is attributed to post-tetanic potentiation (PTP) whereas the smaller but longer lasting increase is attributed to long-term potentiation (LTP). 3. Higher frequencies of tetanic stimulation gave increased PTP and LTP. 4. In order to test whether the influx of calcium ions into the nerve terminal during the tetanus is likely to be involved in potentiation, facilitation was measured during PTP and LTP. Facilitation was reduced to approximately zero during PTP but recovered to normal values about 15 min into LTP. A requirement for the induction of LTP was shown to be the presence of calcium in the bathing solution. However, blocking synaptic transmission with a high concentration of hexamethonium (3 mM) during the tetanic stimulation did not block the induction of LTP. 5. Application of the muscarinic inhibitor, atropine (2 microM), did not affect the magnitude of PTP or LTP.(ABSTRACT TRUNCATED AT 250 WORDS)     

The effect of carbamazepine on the post-tetanic twitch tension in the diaphragm of the mouse

The effects of carbamazepine (0.084-0.25 mM) on the post-tetanic potentiation of the twitch tension, were studied on the isolated phrenic nerve diaphragm preparation of the mouse. Carbamazepine decreased the post-tetanic potentiation of the twitch tension. The maximal depressant effect was found after higher frequencies and longer durations of stimulation. After repetitive stimulation, the amplitude of the endplate potential and the frequency of miniature endplate potentials were potentiated. Carbamazepine decreased the post-tetanic potentiation of the amplitude of endplate potential and the frequency of miniature endplate potentials. The directly-elicited muscle action potential was not affected when post-tetanic potentiation of the twitch tension was decreased. It is concluded that carbamazepine suppressed the post-tetanic potentiation of the indirectly-elicited twitch tension, mainly due to its pre-synaptic inhibitory effect.     

Mechanisms Underlying the Vecuronium-Induced Tetanic Fade in the Isolated Rat Muscle

Abstract The cellular mechanisms underlying the effects of vecuronium on the tetanic contraction were studied in vitro with a combination of myographic and electrophysiologic techniques. We used the isolated sciatic nerve extensor digitorum longus muscle preparation of the rat. Indirect twitches were evoked at 0.1 Hz pulses and tetani at 50 Hz pulses. Trains of end-plate potentials were generated at 50 Hz. The electrophysiological variables used in the analysis of the end-plate potentials were: amplitude, tetanic run-down, quantal size and quantal content. The myographic study demonstrated that vecuronium at 0.4 μM caused tetanic fade, but left the twitch unaffected. Regarding electrophysiology, vecuronium (0.4 μM) decreased the amplitude of end-plate potentials and increased their tetanic run-down. These changes were due to significant reductions in both the quantal content of the end-plate potentials and the quantal size. It is concluded that vecuronium has both pre- and postsynaptic effects at the neuromuscular junction, and that it induces fade of the tetanic contraction via a summation of these effects.     

Enhancement of purinergic neurotransmission by galantamine and other acetylcholinesterase inhibitors in the rat vas deferens

Galantamine, a mild acetylcholinesterase inhibitor and an allosteric ligand of nicotinic receptors, enhanced in a concentration-dependent manner the amplitude of purinergic twitch contractions of the electrically stimulated rat vas deferens (0.2 Hz, 1 ms, 60 V). Other acetylcholinesterase inhibitors also increased the twitches, showing a hierarchy of potencies of galantamine>physostigmine>tacrine>rivastigmine=donepezil. The potentiations seem to be unrelated to the ability to inhibit acetylcholinesterase, since the hierarchy of potencies to block the enzyme in vas deferens was tacrine>physostigmine>rivastigmine>donepezil>galantamine. Acetylcholine also increased the twitches; such effect was produced by a low range of concentrations of acetylcholine (10(-10)-10(-7) M). This facilitatory effect of acetylcholine on twitches was significantly potentiated by galantamine (10(-7)-10(-6) M), but not by rivastigmine or donepezil. A striking enhancement of twitches was also caused by charybdotoxin, a blocker of high-conductance Ca2+-activated K+ channels, and by 4-aminopyridine, a non-specific blocker of K+ channels; in addition, apamin, a blocker of small-conductance Ca2+-activated K+ channels, induced a lower potentiation. The antagonist mecamylamine (10(-7)-10(-6) M) reduced by 80% the potentiation by galantamine, indicating the involvement of nicotinic receptors. Therefore, it is suggested that, besides an inhibition of acetylcholinesterase, some additional mechanisms, such as blockade of Ca2+-dependent K+ channels, or activation of nicotinic receptors of nerve terminals, might be involved in twitch potentiation. These results are relevant in the context of the clinical use of galantamine to improve cognition and behaviour in patients with Alzheimer's disease.     

Effects of memantine on the twitch tension of mouse diaphragm

The effects of memantine (50-175 microM) on the post-tetanic potentiation of the twitch tension were studied on the isolated mouse nerve diaphragm preparation. Memantine completely abolished the twitch tension elicited indirectly while it had no effect on the directly elicited twitch tension. Memantine also decreased the post-tetanic potentiation of amplitude of endplate potential and twitch tension. The duration of tetanic stimulation that induced a maximal decrease of twitch tension was 10-20 s. It is suggested that the effect of memantine on post-tetanic potentiation may be due to its voltage- and time-dependent effect on the ion channel-acetylcholine receptor complex.     

Post-tetanic potentiation at the nerve-muscle junction in the longitudinal muscle of the guinea-pig ileum

The effect of short tetanic stimulation (30 Hz for 25 s) on the following twitch responses of the myenteric plexus-longitudinal muscle preparation of guinea-pig ileum to electric stimulation (0.1 Hz) was investigated in the presence of naloxone and indomethacin. Post-tetanic potentiation (PTP) of the twitches observed in control experiments was abolished in preparations desensitized by substance P but it was not affected in preparations desensitized by serotonin or pretreated with methysergide. Immediately after 5 min tetanic stimulation a decreased sensitivity to substance P but unchanged sensitivity to serotonin were observed. Electromyogram (EMG) of the longitudinal muscle layer was picked up 4 and 10 mm aborally from the stimulation site in response to 1 to 16 impulse trains delivered at 100 Hz. In control conditions only the longer trains triggered this neurogenic response at the distal recording site. In the presence of substance P but not serotonin facilitation occurred so that the distal site was frequently recruited to respond with an EMG even to single impulses. A substance P-like compound rather than serotonin may be a candidate for the neuromodulator or neurotransmitter substance involved in PTP and changes in the response topography of muscarinic transmission.     

Modification by decreased temperature and hypoxia of 45Ca movements in stimulated smooth muscle of rabbit aorta

In rabbit aortic smooth muscle, contractile responsiveness to 10(-6) M norepinephrine or 60 mM added K+ was inhibited by hypoxia (induced by 100% N2) or decreased bath temperature. Hypoxia increased K+-induced 45Ca efflux, inhibited K+-induced 45Ca uptake, and inhibited norepinephrine-induced 45Ca release; lowering bath temperature by 10 degrees C decreased resting 45Ca uptake and norepinephrine-induced 45Ca release by 30-40% and decreased the 45Ca uptake elicited with norepinephrine or K+ by more than two-thirds. Thus, hypoxia and temperature variations affect different Ca2+ components. Hypoxia decreases the norepinephrine-sensitive membrane Ca2+ fraction and the K+-stimulated mitochondrial Ca2+ fraction whereas decreased temperature most strongly inhibits membrane-associated Ca2+ uptake increases elicited with either norepinephrine or high K+.     

Effects of modulating Ca2+ entry and activating prejunctional receptors on facilitation of excitatory junction potentials in the guinea-pig vas deferens in vitro

This study investigated the effects of changing the extracellular Ca2+ concentration on facilitation of excitatory junction potential (EJP) amplitude during trains of 20 stimuli at 1 Hz at sympathetic neuroeffector junctions in the guinea-pig vas deferens. These effects were compared with those of Ca2+ channel blockers and agents which act at prejunctional receptors to increase or decrease neurotransmitter release. In these experiments, alpha-adrenoceptor-mediated autoinhibition of neurotransmitter release was blocked by the alpha-adrenoceptor antagonist, phentolamine (1 microM). Varying the extracellular Ca2+ concentration (0.75-6 mM) changed the amplitude of EJPs without affecting the maximum level of facilitation during the trains of stimuli. Reductions in Ca2+ concentration (from 2 mM) were associated with a slowing in the rate of development of facilitation. The Ca2+ channel antagonists, Cd2+ (2 microM and 5 microM) and omega-conotoxin GVIA (10 nM), and agents which act at prejunctional receptors to reduce neurotransmitter release, adenosine (100 microM and 1,000 microM) and prostaglandin E2 (PGE2; 0.1 nM and 1 nM), produced similar effects to those of lowering the extracellular Ca2+ concentration. Raising the extracellular Ca2+ concentration (from 2 mM) increased the rate of development of facilitation. Angiotensin II (AII; 0.5 microM) produced similar effects to raising extracellular Ca2+. However, isoprenaline (1 microM), while increasing EJP amplitude, reduced the maximum level of facilitation and was without effect on the rate of development of facilitation. In the guinea-pig vas deferens EJPs are produced by neurally released ATP. Thus, the findings support the idea that adenosine, PGE2 and AII change ATP release by modifying Ca2+ entry into the nerve terminal. However, the effects of isoprenaline may not solely be accounted for by modifications in Ca2+ entry.