Remy S. Leeuwin

The effect of corticosteroids and hemicholinium-3 on choline uptake and incorporation into acetylcholine in rat diaphragm

The glucocorticoids prednisolone and dexamethasone antagonize the inhibition by hemicholinium-3 of both the rate of choline uptake and the incorporation of choline into acetylcholine in the rat diaphragm. Aldosterone has no such effects. It is concluded that the beneficial effect of glucocorticoids in the treatment of myasthenia gravis may be due not only to immunosuppression, but also to some direct effect on presynaptic events perhaps via a choline carrier or an enzyme of choline metabolism.

Effect of corticosteroids on sciatic nerve‐tibialis anterior muscle of rats treated with hemicholinium‐3 An experimental approach to a possible mechanism of action of corticosteroids in myasthenia gravis

We studied the effect of intraperitoneally administered corticosteroids on the neuromuscular transmission in the sciatic nerve-tibialis anterior muscle preparation of the anesthetized rat stimulated at a rate of 10 Hz. Administered simultaneously with hemicholinium-3 (HC-3), 80 pg per kilogram (that is, half the lethal dose for 50 percent survival), prednisolone and dexamethasone cause a marked reversal of the block of the neuromuscular transmission caused by HC-3. The effect of aldosterone is very small. The blocking action of d-tubocurarine is not antagonized by either prednisolone or dexamethasone. Choline provides total protection against the HC-3 blockade, whereas physostigmine, in a just sublethal dose, is ineffective. We tentatively conclude that in myasthenia gravis the carrier-mediated transport of choline into the nerve endings may be deficient and that the beneficial effect of corticosteroids in this condition is based on their ability to ameliorate the deficient choline transoort.

Effect of corticosteroids on the phrenic nerve‐diaphragm preparatior treated with hemicholinium A possible model of myasthenia gravis

A marked protective action of the corticosteroids prednisolone, hydrocortisone, and dexamethasone has been shown on a hypothetical model of myasthenia gravis, using the rat phrenic nerve-diaphragm preparation treated with hemicholinium-3. Dexamethasone is much more effective than prednisolone, and hydrocortisone is the least effective. Prednisolone has no effect on a neuromuscular block caused by choline, decamethonium, physostigmine, d-tubocurarine, and a high or low calcium ion concentration. The possible implications of the present study for myasthenic disease are discussed. It is tentatively concluded that the site of action of corticosteroids in myasthenia gravis is located presynaptically.A marked protective action of the corticosteroids prednisolone, hydrocortisone, and dexamethasone has been shown on a hypothetical model of myasthenia gravis, using the rat phrenic nerve-diaphragm preparation treated with hemicholinium-3. Dexamethasone is much more effective than prednisolone, and hydrocortisone is the least effective. Prednisolone has no effect on a neuromuscular block caused by choline, decamethonium, physostigmine, d-tubocurarine, and a high or low calcium ion concentration. The possible implications of the present study for myasthenic disease are discussed. It is tentatively concluded that the site of action of corticosteroids in myasthenia gravis is located presynaptically.

Effects of corticosteroids on neuromuscular blocking actions of d-tubocurarine

Dexamethasone (50 microgram/kg) significantly increased the LD50 of d-tubocurarine (d-TC) when administered i.p. simultaneously with d-TC. Choline (50 and 100 mg/kg) gave some protection against the lethal effects of d-TC and the cholinesterase inhibitors neostigmine (250 microgram/kg) and physostigmine (1000 microgram/kg) provided full protection against doses of d-TC twice the LD50. The blocking effect of d-TC (75 microgram/kg) on the sciatic nerve-tibialis anterior muscle preparation was antagonized by dexamethasone. Prednisolone delayed the occurrence of a complete neuromuscular block caused by d-TC in the phrenic nerve-diaphragm preparation, and antagonized the effect of d-TC on short tetanic contractions. d-TC (5 mumol/l) inhibited the [14C]choline uptake in the endplate-rich region of the rat diaphragm during stimulation. This inhibition was antagonized by dexamethasone as well as by physostigmine. The incorporation of radioactive choline into acetylcholine was inhibited in the presence of d-TC (15 mumol/l), and both dexamethasone and physostigmine counteracted this inhibition. It is concluded from these experiments that d-TC very probably has an effect on the choline carrier system. These experimental results support the hypothesis that glucocorticoids may improve reduced muscle performance by direct presynaptic effects at the neuromuscular junction.

Interactions of cholinesterase inhibitors and glucocorticoids with ketamine and pentobarbitone-induced general anaesthesia in the rat: possible effects on central cholinergic activity

1 Doses of 100, 150 and 200 μg kg−1 of the Cholinesterase inhibitor neostigmine reverse the anaesthetic action of ketamine. The antagonistic effect is increased as the dose is increased. The duration of anaesthesia induced by pentobarbitone is reversed by the Cholinesterase inhibitor in doses of 150, 200 and 250 μg kg−1. 2 Choline, in a dose of 50 mg kg−1, significantly antagonizes the action of the two anaesthetics, whereas hemicholinium‐3, an inhibitor of the uptake of choline and the synthesis of acetylcholine, markedly potentiates their action. 3 Dexamethasone induces a significant reduction of the duration of anaesthesia produced by ketamine and pentobarbitone. The potentiation of the anaesthetic effect caused by hemicholinium‐3 is also reversed by dexamethasone. 4 The acetylcholine content in rat cerebral cortex is increased after treatment with ketamine and pentobarbitone. 5 Measurements of the course of the plasma level of pentobarbitone do not reveal alterations in the pharmacokinetic profile by either neostigmine or dexamethasone. 6 These results indicate that central cholinergic systems may somehow be involved in the anaesthesia induced by ketamine and pentobarbitone and that the interactions described in this paper may be the result of modification by neostigmine and dexamethasone of the alterations in cholinergic activity caused by the two anaesthetics.

The effect of prednisolone on the rat phrenic nerve-diaphragm preparation treated with hemicholinium

Abstract Recent papers have reported the benificial action of corticosteroids in myasthenia gravis. The underlying mechanism of action is not known. In a preliminary investigation the effect of prednisolone on the rat phrenic nerve-diaphragm preparation was studied. It was shown that a relatively high stimulation rate which caused ‘fatigue’ of the preparation, prednisolone prevented the blocking effect of hemicholinium-3, but had no effect on the preparation prior to addition of hemicholinium.

Antagonism of corticosteroids against the lethal effects of hemicholinium-3 in rats and mice

The corticosteroids prednisolone or dexamethasone administered simultaneously with hemicholinium-3 (HC-3) by i.p. injection, increased the LD50 of HC-3 in rats and mice about twofold. Daily treatment with prednisolone for 7 days also increased the LD50 of HC-3 in rats. Choline completely protected rats and mice against HC-3. A sublethal dose of physostigmine did not change the LD50 of HC-3 in rats but increased the LD50 in mice to a small extent. The possible implications of these experiments for the explanation of the beneficial effects of corticosteroids in myasthenia gravis are discussed.

The effect of cholinesterase inhibitors and corticosteroids on rat nerve-muscle preparations treated with hemicholinium-3

Corticosteroids improve muscle function impaired by hemicholinium-3 (HC-3). The effects of cholinesterase inhibitors either alone or in combination with the glucocorticoid dexamethasone were studied on nerve-muscle preparations treated with HC-3, both in vivo and in vitro. Administered simultaneously with HC-3 (80 μg/kg by i.p. injection, physostigmine (250–1500 μg/kg) caused a significant, dose-dependent reversal of the neuro-muscular transmission block caused by HC-3 in the sciatic nerve-tibialis anterior muscle preparation of the anaesthetized rat, stimulated at a rate of 10 Hz. The HC-3-blocked neuromuscular transmission was also antagonized by neostigmine (125 and 250 μg/kg). Under the same conditions physostigmine (1000 μg/kg) potentiated the effect of dexamethasone. After i.p. injection of physostigmine (125–1000 μg/kg) and neostigmine (62.5–250 μ/kg) respectively, the LD50 in rats was significantly increased. With physostigmine a sharp maximum was found at 250 μg/kg. Neostigmine, and to a lesser extent physostigmine antagonized the neuromuscular transmission inhibited by HC-3 in the in vitro phrenic nerve—diaphragm preparation of the rat. The HC-3-induced inhibition of both uptake of choline and its incorporation into acetylcholine was antagonized by physostigmine and neostigmine. This antagonism was not altered in the presence of dexamethasone 0.2 μM. The possible implications of these findings, and the mechanism underlying the effects seen with corticosteroids and cholinesterase inhibitors are discussed. It is concluded that under certain circumstances a nerve-muscle preparation treated with HC-3 may be an adequate model for myasthenia gravis.

PK 11195 antagonizes the positive inotropic response of the isolated rat heart to diazepam but not the negative inotropic response

The influence of the ligand PK 11195 (1-(2-chlorophenyl)-N-methyl-N-(1- methylpropyl)-3-isoquinolinecarboxamide), antagonist of the peripheral-type benzodiazepine receptor, on the inotropic response of the perfused rat heart to diazepam (7-chloro-5-phenyl-methyl- 1,3-dihydro-2H-1,4-benzodiazepin-2-one) was studied. Diazepam induced a positive inotropic response which was preceded by a transient negative inotropic response. Concentrations of 10(-7) M PK 11195 were ineffective, whereas concentrations of 10(-6) and 10(-5) M PK 11195 reduced the positive inotropic response significantly. At 5 x 10(-5) M PK 11195 the response was completely abolished. The negative inotropic response was not changed by either concentration of PK 11195 used. It is concluded that the positive inotropic response of the isolated rat heart to diazepam may well be mediated through peripheral-type benzodiazepine receptors; the negative inotropic response must be related to other (more complex) mechanisms.

Modification of cardiac actions of Ro 05-4864 by PK 11195 and flumazenil in the perfused rat heart

The benzodiazepine analogue Ro 05-4864 [chlorodiazepam] (2.10[-5] to 4.10[-4] M) induced a concentration-dependent increase of coronary flow rate (Emax 82.4% [+/- 2.2 SEM]) and an increase of contraction force (Emax 68.3% [+/- 4.7 SEM]) of the retrograde perfused, isolated Langendorff rat heart. The influence of PK 11195, antagonist of the peripheral type benzodiazepine receptor, and flumazenil (Anexate), antagonist of the central type benzodiazepine receptor, on these responses to Ro 5-4864 was studied. In concentrations of 10(-7) to 5.10(-5) M, PK 11195 significantly reduced both the increase of coronary flow rate and of contraction force, without affecting these functions by itself; the positive inotropic response produced by Ro 05-4864 was even abolished in the presence of 5.10(-5) M PK 11195. The Emax values of Ro 05-4864 on both coronary flow and inotropy were reduced significantly by PK 11195. In the presence of flumazenil, 10(-7) to 10(-5) M, both the vasodilatory and the positive inotropic response induced by Ro 05-4864 were significantly counteracted as well. The Emax values of Ro 05-4864 were reduced significantly. In conclusion, the results support earlier suggestions that it is tempting to involve peripheral type benzodiazepine receptors in cardiac actions of benzodiazepines. The finding that the centrally acting benzodiazepine antagonist flumazenil reduced the cardiac actions of Ro 05-4864 is as yet difficult to explain. On the other hand concentrations of both agonist and antagonist employed are so-much high that interference of other receptors than benzodiazepine receptors must be considered as well.