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.     

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.     

Effect of neomycin on post-tetanic twitch tension of the mouse diaphragm preparation.

The effect of calcium (0.5-6 mM) and neomycin (0.1-0.2 mM) on the maximum post-tetanic twitch tension (MTT) and post-tetanic depression (PTD) of the indirectly elicited twitch tension was studied on the mouse isolated phrenic nerve-diaphragm preparation. The effect of neomycin on MTT of directly stimulated twitch tension was also tested in (+)-tubocurarine pretreated preparations. Three-dimensional plots between MTT and frequency and duration of indirect tetanic stimulation revealed that the frequencies and durations inducing maximal MTT were 500 Hz for 20 s in 0.5 mM CaCl2, 100 Hz for 5 s in 2 mM CaCl2 and 100 Hz for 10 s in 6 mM CaCl2. The frequency and duration inducing maximal PTD was 100 Hz for 20 s in 0.5 mM CaCl2, but there was no PTD in 2 mM or 6 mM CaCl2. Neomycin was associated with significantly greater MTT than in control if the duration of tetanic stimulation was 1 or 2 s, while it was associated with less MTT if the duration of tetanic stimulation was 10 or 20 s. Neomycin caused PTD in 2 mM CaCl2; sometimes the depressive effect was so severe that twitch tension was abolished. The maximal depression effect was found after 100 Hz tetanic stimulation for 20 s. Increasing the extracellular calcium concentration to 6 mM antagonized the effects of neomycin on MTT and PTD, whereas neostigmine (1.6 microM) antagonized the effect partially. Neomycin had no effect on the MTT or PTD of the directly stimulated twitch tension. It is concluded that neomycin alters the conditions of tetanic stimulation inducing MTT.     

Effects of hypoxia, 2,4-dinitrophenol and ouabain on twitch potentiation elicited following conditioning pulses in the mouse phrenic nerve-diaphragm preparation

Hypoxia is suggested to increase intracellular Ca2+, thereby affecting cellular functions. If this is the case, the effect would be modified by other treatments which also elevate intracellular levels of Ca2+. To test this possibility, in mouse diaphragm preparations, the effects of hypoxia were examined on twitch potentiations after application of neural conditioning pulses which are considered to elevate intracellular Ca2+. The effects were compared with those of 2,4-dinitrophenol (DNP) and ouabain. Hypoxia increased the tension of twitches elicited directly or neurally with 0.1 or 0.5 Hz pulses at 36 degrees C but not at 24 degrees C. Paired pulses to the nerve induced a complex response profile in which the amplitude of the second twitch was enhanced (twitch facilitation). Tetanic pulses (50 Hz) to the nerve were followed by post-tetanic twitches of increased amplitude (post-tetanic twitch potentiation, PTP). Hypoxia little affected the twitch facilitation but abolished PTP at both temperatures. These effects differed from those of DNP and ouabain in some ways. Thus, present experiments indicate that hypoxia selectively influences the process which is responsible for the PTP phenomenon rather than for the twitch facilitation. It is possible that the mechanism by which hypoxia would accumulate intracellular Ca2+ may be included in the process through which PTP occurs.     

Dantrolene effects on neuromuscular function in cat soleus muscle.

Dantrolene sodium (DS) was investigated for its effects on cat soleus muscle contractile properties and motor nerve terminal activity in particular. DS, 0.1-1.5 mg/kg i.v., caused a dose-dependent depression of indirectly elicited contractile strength which was more pronounced at lower frequencies of stimulation. Maximum tetanic strength at frequencies of 10-400 Hz was depressed to a lesser degree than contractile responses evoked by lower frequencies of stimulation; the twitch/tetanus contraction ratios were reduced with increasing dose, primarily because of diminished twitch. DS was without effect on motor nerve terminals as evidenced by normal post-tetanic repetition in the nerves following DS administration. Post-tetanic potentiation became relatively larger in amplitude as contractile strength was diminished. These data suggest that DS depresses neuromuscular function at a site other than the neural apparatus at the neuromuscular junction.     

A comparison between the effects of a tetanus and the effects of sympathomimetic amines on fast- and slow-contracting mammalian muscles

The effects of adrenaline and isoprenaline on the tension and time-course of the contractions of the tibialis anterior and soleus muscles of cats and rabbits have been compared with the effects of previous high-frequency stimulation. Like a tetanus, adrenaline possessed a facilitating action on neuromuscular transmission and an action exerted directly on the muscle fibres. Isoprenaline possessed only the second of these two actions. The effect of adrenaline and isoprenaline on the muscle fibres was blocked by dichloroisoprenaline and by 1-(2 naphthyl)-2-isopropylaminoethanol, whereas the effect of adrenaline on neuromuscular transmission was blocked by phentolamine and by phenoxybenzamine. In the soleus muscle, both the catechol amines and a previous tetanus caused similar decreases in maximal twitch tension and in the times to peak tension and to half-relaxation. The muscle action potentials were unaltered or slightly increased in amplitude. In the tibialis anterior muscle, a previous tetanus and the catechol amines caused an increase in twitch tension and an increase in the overall duration of the twitch. The muscle action potentials were either unchanged or were slightly decreased in amplitude. In this muscle the effect of a tetanus differed from that of the catechol amines in that the large post-tetanic change was associated with a marked increase in the rate of rise of twitch tension.     

In vitro galantamine-memantine co-application: mechanism of beneficial action

Several drugs are in clinical use for symptomatic treatment of Alzheimer's disease patients. Since Alzheimer's disease is known to be associated with down-regulation of the cholinergic and N-methyl-d-aspartate (NMDA) systems, most of these drugs inhibit acetylcholinesterase, potentiate the activity of nicotinic acetylcholine receptors (nAChRs), or modulate NMDA receptors. Galantamine is an anticholinesterase and allosterically potentiates the activity of the nicotinic receptors. We have recently found that galantamine potentiates the activity of NMDA receptors as well. Memantine is unique in that it inhibits the NMDA receptors. We have developed a hypothesis that combining galantamine and memantine will be more effective for improving the patient's conditions than monotherapy with either drug. Patch clamp and intracellular Ca²⁺ imaging experiments using rat cortical and hippocampal neurons clearly provided the in vitro bases for our hypothesis. Memantine blocked the extrasynaptic NMDA receptor 100 times more potently than the synaptic NMDA receptor at negative membrane potentials and the block of both types of NMDA receptors was attenuated with depolarization. However, galantamine potentiation of the NMDA receptors was not voltage dependent. Thus, co-application of memantine with galantamine prevented the galantamine potentiation and the activation of extrasynaptic NMDA receptors, but membrane depolarization revealed the galantamine potentiation. Therefore, cell death is expected to be prevented by memantine near the resting potential while the NMDA-mediated synaptic transmission, which is down-regulated in the patients, is maintained and potentiated by galantamine. These results provide in vitro bases for the beneficial actions of galantamine and memantine combinations.     

Blocking effect of diltiazem in the isolated rat phrenic hemidiaphragm

The effects of diltiazem on indirectly and directly elicited twitch were studied in the isolated rat phrenic hemidiaphragm preparation. Diltiazem (30-500 microM) blocked the indirectly elicited twitch response and this effect was not affected by reducing the extracellular calcium from 2.5 to 1.25 mM. An effect on the directly elicited twitch was also observed (100-300 microM). Diltiazem (30-300 microM) blocked the peak tetanic tension and tetanic fade was present. The results were consistent with an action of diltiazem on the nicotinic receptor-ion channel complex.     

Memantine potentiates hippocampal theta oscillations at a therapeutic dose in anesthetized mice: a mechanistic link to its cognitive-enhancing properties

Memantine is an uncompetitive, low-affinity NMDA receptor antagonist clinically used for the treatment of cognitive deficits in moderate to severe Alzheimer's disease. Both neurophysiological and behavioral studies in rodents have suggested a beneficial effect of memantine on synaptic plasticity and learning performances. In the present study, we investigated the effect of memantine on pedonculopontine-elicited theta oscillations in the hippocampus of urethane anesthetized mice, a model shown to be sensitive to several pharmacological agents exhibiting cognitive-enhancing properties. We found that a low dose of memantine potentiated elicited theta power while a high dose was disruptive. The low dose of memantine used was shown to yield an unbound brain concentration well within the range of therapeutic concentrations reported in rodent brain extracellular fluid and human cerebrospinal fluid. For further comparison, the effect of another uncompetitive NMDA receptor antagonist with higher affinity, i.e. MK-801, was also investigated. MK-801 was at a low dose devoid of effect on elicited theta power, while a high dose, within the range of doses reported to induce cognitive deficits in a variety of hippocampal-dependent learning paradigms in mice, was found disruptive on elicited theta waves. Taken together, our results suggest that clinically relevant doses of memantine promote neuronal network synchronization in the hippocampus, which may represent an underlying mechanism for the reported cognitive-enhancing properties in both preclinical and clinical studies.     

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.     

Reduction by diazepam of repetitive electrical activity and toxicity resulting from soman

The organophosphate cholinesterase inhibitor soman (methyl pinacolylphosphufluoridate) produces potentiation of twitch tension in rat anterior tibial and phrenic nerve-diaphragm preparations by initiating repetitive electrical activity in the vicinity of the motor nerve terminals. Diazepam abolished this repetitive electrical activity and hence the associated potentiation of muscle twitch tension. These effects are produced by concentrations of iazepam that are without other apparent effects on neuromuscular transmission, as indicated by the absence of effect on normal twitch responses. The combination of diazepam with atropine provides additional protection against soman toxicity in rabbits.     

Effects of memantine and MK-801 on NMDA-induced currents in cultured neurones and on synaptic transmission and LTP in area CA1 of rat hippocampal slices.

The effects of the uncompetitive N-methyl-D-aspartate (NMDA) receptor antagonists, memantine (1-amino-3,5-dimethyladamantane) and MK-801 ((+)-5-methyl-10,11-dihydro-5H-dibenzocyclo-hepten-5,10-imin e maleate) were compared on synaptic transmission and long-term potentiation (LTP) in hippocampal slices and on NMDA-induced currents in cultured superior collicular neurones. 2. Memantine (10-100 microM) reversibly reduced, but did not abolish, NMDA receptor-mediated secondary population spikes recorded in area CA1 of hippocampal slices bathed in Mg(2+)-free artificial cerebrospinal fluid. 3. Memantine (100 microM) antagonized NMDA receptor-mediated excitatory postsynaptic currents recorded in area CA1 in a strongly voltage-dependent manner i.e. depressed to 11 +/- 4% of control at -35 mV and 95 +/- 5% of control at +40 mV (n = 9), with no apparent effect on response kinetics. 4. The effects of MK-801 and memantine on the induction of LTP were assessed after prolonged pre-incubations with these antagonists. When present for 6.6 +/- 0.4 h prior to tetanic stimulation, memantine blocked the induction of LTP with an IC50 of 11.6 +/- 0.53 microM. By comparison, similar long pre-incubations with MK-801 (6.4 +/- 0.4 h) blocked the induction of LTP with an IC50 of 0.13 +/- 0.02 microM. 5. Memantine and MK-801 reduced NMDA-induced currents in cultured superior colliculus neurones recorded at -70 mV with IC50s of 2.2 +/- 0.2 microM and 0.14 +/- 0.04 microM respectively. The effects of memantine were highly voltage-dependent and behaved as though the affinity decreased epsilon fold per 50 mV of depolarization (apparent delta = 0.71). In contrast, under the conditions used, MK-801 appeared to be much less voltage-dependent i.e. affinity decreased epsilon fold per 329 mV of depolarization (apparent delta = 0.15). 6. Depolarizing steps from -70 mV to +50 mV in the continuous presence of memantine (10 microM) caused a rapid relief of blockade of NMDA-induced currents from 83.7 +/- 1.9% to 21.8 +/- 1.8% (n = 5). This relief was best fitted by a double exponential function (17.2 +/- 11.7 and 698 +/- 204 ms), the faster component of which was most pronounced. 7. In conclusion, whereas MK-801 is equipotent in blocking NMDA-induced currents (at - 70 mV) and the induction of LTP, memantine is relatively less potent in blocking the induction of LTP. This is due to its rapid relief of blockade upon depolarization; a property which might explain its promising clinical profile in the treatment of chronic neurodegenerative diseases.     

Effects of memantine, a non-competitive N-methyl-D-aspartate receptor antagonist, on genomic stability

Memantine is an aminoadamantane drug useful in neurodegenerative diseases, with beneficial effects on cognitive functions. Some studies have shown that memantine protects brain cells, thereby decreasing glutamate excitotoxicity. This study evaluated the genotoxic/antigenotoxic and mutagenic effects of memantine in CF-1 mice, following standardized protocols. Memantine was administered i.p. at 7.5, 15 or 30 mg/kg for three consecutive days. Blood and brain samples were collected to assess DNA damage using the alkaline comet assay. The mutagenic effect was assessed using the bone marrow micronucleus test. In addition, possible antioxidant effects were evaluated measuring the survival of Saccharomyces cerevisiae yeast strains [wild-type (WT) and isogenic mutants lacking superoxide dismutase] to cotreatment of memantine plus hydrogen peroxide. Memantine decreased DNA oxidative damage mainly in brain tissue. This antigenotoxic effect corroborated an increase observed in the survival of S. cerevisiae WT strain against hydrogen peroxide-induced damage. Furthermore, memantine did not increase the micronucleus frequency. The overall results indicate that memantine showed no mutagenic activity, did not cause DNA damage in the blood and brain tissues and showed antigenotoxic effects in brain tissue.     

Effects of isoprenaline on contractions of directly stimulated fast and slow skeletal muscles of the guinea-pig

1. The actions of isoprenaline on the contraction and the resting potential of the isolated extensor digitorum longus (EDL), a fast contracting muscle, and the soleus, a slow contracting muscle, of the guinea-pig were investigated. Twitch tension was elicited by direct supramaximal stimulation and recorded isometrically.2. The twitch tension of EDL elicited by pulses of 0.5-10 ms duration was increased in the presence of isoprenaline (1 mug/ml). Isoprenaline increased the twitch tension of the soleus elicited by a pulse of more than 5 ms duration, but decreased it when elicited by a pulse of less than 1 millisecond. These effects were blocked by propranolol (1-3 mug/ml) but not by phentolamine (1-5 mug/ml).3. In EDL, isoprenaline prolonged the time to peak tension and the half-relaxation time. The twitch of the soleus was shortened by isoprenaline due to an acceleration of relaxation. These findings were independent of stimulus duration.4. The potentiating effects of isoprenaline on the twitch tension of EDL and the soleus were not observed in K(+)-free Krebs solution and were abolished by ouabain (1 mug/ml) and by reduction of the temperature from 33 degrees to 18 degrees C. The effects of isoprenaline on the relaxation proces were not affected by these treatments.5. In EDL, the resting potential increased from 77.3 mV to 78.5 mV after isoprenaline, whereas in the soleus it increased from 69.1 to 74.7 mV. These effects were blocked by propranolol, K(+)-deficiency, ouabain, and cooling to 18 degrees C. Hyperpolarization by isoprenaline was increased by substitution of isethionate for the external chloride.6. There was a good correlation between the potentiation of the mechanical response and the hyperpolarization of the membrane by isoprenaline. The hyperpolarization seems to be due to activation of the Na(+)-K(+) pump.     

Post-tetanic potentiation in the innervated smooth muscle preparation of the guinea-pig ileum

The phenomenon of posttetanic potentiation of electrically evoked twitch contractions in the myenteric plexus-longitudinal muscle of the guinea-pig ileum was observed when (1) post-tetanic inhibition had been prevented by naloxone, an opiate receptor antagonist, and (2) the twitches had been diminished by indomethacin, an inhibitor of prostaglandin synthesis. The origin of posttetanic potentiation at this muscarinic synapse appeared to be presynaptic.     

Memantine upregulates BDNF and prevents dopamine deficits in SIV-infected macaques: a novel pharmacological action of memantine.

N-methyl-D-aspartate (NMDA) receptor activation is involved in the pathogenetic cascades of neurodegenerative disorders including human immunodeficiency virus (HIV) dementia. Memantine, an uncompetitive NMDA receptor antagonist, which has been recently approved for the treatment of Alzheimer's disease, is being discussed as a potential adjunctive therapeutic substance for HIV dementia. We used simian immunodeficiency virus-infected rhesus macaques to assess the effects of memantine on brain dysfunction and brain pathology within 3-5 months after initial infection during early asymptomatic stage of disease. We had shown previously that within this time frame, marked changes were evident in the dopaminergic systems. Memantine was administered two weeks post infection, at peak viremia, in order to prevent early NMDA receptor activation due to immune mediators. We found that memantine prevented onset of dopamine deficits in the brains of SIV-infected macaques, without affecting early brain pathology or peripheral course of infection. Memantine specifically upregulated mRNA and protein expression of the neurotrophic factor brain-derived neurotrophic factor (BDNF), suggesting that the protective effect of memantine on dopamine function may be mechanistically remote from NMDA receptor antagonism. This novel pharmacological action of memantine may also be relevant for other neurodegenerative disorders and supports the involvement of neurotrophic factors in adult brain neuroprotection.     

Effects of velnacrine, tacrine and physostigmine on tetanic twitch responses at the rat neuromuscular junction.

The effects of velnacrine (1-hydroxytacrine), tacrine and physostigmine on indirectly elicited twitch at low and high stimulation frequencies were analyzed in the rat phrenic hemidiaphragm preparation. At 0.2 Hz, velnacrine and physostigmine behaved in a similar manner, the latter showing a higher potentiating effect. This potentiation was observed at 3-100 microM velnacrine, whereas a slight depression appeared at higher concentrations. When tetanic responses were studied, the drug concentrations needed to depress tetanic tension and tetanic fade were quite different in the case of velnacrine (depression of tetanic tension from 1 microM and tetanic fade from 170 microM), whereas physostigmine and tacrine were able to affect these parameters at very similar concentrations. The results suggest that some effects of velnacrine could differ from those of tacrine in spite of the chemical similarity.     

The effect of memantine on various neurobiological processes

Experiments are reported which show that 1,3-dimethyl-5-aminoadamantane (DMAA, D-145, memantine, Memantine) does not influence the binding capacity of a variety of tritiated ligands to rat brain membranes thus suggesting that the drug does not interact with dopamine, opioid, GABA, alpha 1- and alpha 2-adrenergic receptors, but can influence with low potency 5-HT1 receptors. Autoradiographical studies also show that memantine does not interfere with the specific uptake mechanism of various transmitter substances nor is 3H-memantine taken up by any neuronal elements. Memantine at high concentrations was shown to release 3H-monoamines which had previously been taken up by nervous tissue but the release process is independent of calcium ions and is not dose-dependent. The latter effect could be explained by CNS culture studies which show memantine to produce morphological changes in neurones and glia when present at concentrations greater than 50 mumol/l.     

Effect of some benzodiazepines on peripheral neuromuscular function in the rat in-vitro hemidiaphragm preparation

The effect of equimolar cumulative concentrations of 11 different benzodiazepines on the indirectly evoked twitch contraction was investigated in the rat in-vitro phrenic nerve-hemidiaphragm preparation. Depending on the pattern of the concentration-response curves two groups of benzodiazepines can be distinguished: (i) a first group with a biphasic action, e.g. potentiation of twitch tension in low concentrations and depression of twitch tension in high concentrations, and (ii) a second group with primary depression of twitch tension with increasing concentrations. All of the tested compounds ultimately caused a 100% depression of twitch tension at concentrations ranging from 0.175 to 0.35 mmol litre-1. Although this peripheral effect of benzodiazepines on neuromuscular function is not the main site of action of these compounds, there are enough arguments to state that it is not a simple toxic effect. There is some evidence from this study that the peripheral component of the benzodiazepine effect on muscle relaxation may involve a multi- rather than one single receptor system.