The contraction of intestinal myocells and block of cationic pumps

The effects of a block of Na-K-ATPase pump on the contraction of intestinal smooth muscle (ileum of guinea pig) induced by carbachol has been analysed. Two parameters have been evaluated: a) maximum velocity of contraction (Vmax.C); b) maximum velocity of relaxation (Vmax.D) after washing. The block of pump is produced by a digitalic agent (desacetyl-lanatoside C). The application of 0.3 micrograms/ml of lanatoside for 1 min causes an increase of Vmax.C and a decrease of Vmax.D; whereas it does not influence the height of contraction. This effect is in agreement with the mechanism of action of Na on the contraction curve.     

In vitro study of rat uterus after chronic formaldehyde exposure.

To measure cholinergic, adrenergic and tryptaminergic receptor activity of formaldehyde (HCHO) in rat uterus, albino rats were treated with 5 and 10 mg/kg, ip HCHO for 30 days. Acetylcholine (ACh) in doses 1.33, 2 and 3 micrograms/ml produced mild to moderate contraction of isolated rat uterus in control group. HCHO had no effect on isolated rat uterus per se, however it reduced ACh and carbachol induced contraction and presence of adrenaline influences in respect of ACh and carbachol activity. Adrenaline per se had no effect in control preparations, but reduced carbachol induced contraction. Propranolol had no effect on rat uterus; but its presence in the bathing medium increased activity of adrenaline. 5-Hydroxytryptamine (5-HT) had no effect of its own on isolated rat uterus but its presence in the bathing medium enhanced contractions of carbachol and oxytocin.     

Changes in the activity of acetylcholinesterase and Na,K-ATPase in human erythrocytes irradiated with X-rays

The response of human erythrocytes to X-rays in the dose range from 40 Gy to 600 Gy was determined on the basis of changes in the activities of AChE and ATPase. The Na,K-ATPase activity increased above the control value at doses below 200 Gy, while at the doses higher than 200 Gy, it decreased, reaching 96% of the control value at a dose of 600 Gy. In the range of doses up to 200 Gy, the AChE activity, expressed as Vmax, did not change. At higher doses, it fell drastically, reaching 33% of the control value at a dose of 600 Gy. Simultaneously, the enzyme substrate affinity decreased at 200 Gy, and then started to increase at lower values of Vmax. The obtained results suggest that under appropriate conditions, low doses of radiation may have the opposite effects to high doses.     

Effects of dopaminergic and cholinergic interactions on rat behavior.

The present work studied the effects of dopaminergic and muscarinic receptor agonists and antagonists on rat locomotor activity and catalepsy. Results showed that carbachol at the highest dose used (10 mg/kg, p.o.) decreased and pimozide at the dose used abolished locomotor activity. Atropine at a low dose (1 mg/kg, p.o.) increased and at a high dose decreased this parameter. Mazindol at a high dose also increased locomotor activity. A significant and dose-dependent increase in the time on the bar was observed in animals treated with carbachol or pimozide as compared to controls. The increase observed with pimozide was greater than 60 s. Effects of carbachol on locomotor activity were observed already after the first drug exposure, but the increased time on bar produced by this drug in the test of catalepsy was observed only after repeated exposure (7th day). The effect of the highest dose (10 mg/kg, p.o.) of atropine (decreased activity) as related to the lowest one was evident at the 7th day, but the increased locomotor activity seen at the low dose was detected already at the first day. There was a predominance of the effect of pimozide on the open field as well as on catalepsy after its association with each one of the three doses of carbachol. The association of atropine and mazindol did not seem to alter locomotor activity and catalepsy as related to each drug alone. Our results indicate that interactions between dopaminergic and cholinergic systems play an important role on behavior and motor functions.     

Effects of liver blood flow on hepatic uptake kinetics of galactose in anesthetized cats: parallel tube model

Hepatic galactose uptake in cats anesthetized with pentobarbital was determined during (i) steady-state infusions at several doses, (ii) rapidly increasing infusion rates at different blood flows, and (iii) prolonged infusion of a single dose at different blood flows. The hepatic venous long-circuit technique was used to allow frequent sampling of arterial, portal, and hepatic venous blood without depletion of the animal's blood volume and to allow measurement and alteration of total hepatic blood flow. Uptake was shown to follow Michaelis-Menten kinetics and was consistent with the "parallel tube model." The kinetic parameters Vmax and Km could be determined under steady-state and nonsteady-state conditions and were independent of hepatic blood flow over the range 60-150% of control flow. Mean Vmax was 80 mumol/(min X 100 g liver) and mean Km was 215 microM. Vmax declined by 50% when flow was reduced to half normal. It is concluded that the parallel tube model can be used to describe and predict hepatic galactose kinetics in anesthetized cats, although other models may fit the data equally well.     

In vitro tracheal responses from mice chosen for in vivo lung cholinergic sensitivity

We selected two inbred strains of mice based on their different in vivo lung responses to intravenous acetylcholine for studies on the in vitro tracheal responses to contractile and relaxing agents. In addition, we studied the role of cyclooxygenase products on the in vitro responses. Tracheal rings were contracted with increasing concentrations of carbachol and KCl and relaxed with increasing concentrations of isoproterenol after contraction with carbachol at the concentration that produced 30, 50, and 70% of the maximal contraction (EC30, EC50, and EC70, respectively) and KCl at the EC50. Half the tracheae simultaneously underwent the same protocols after pretreatment with indomethacin (3 X 10(-6) M). Despite a severalfold difference in the maximal response to cholinergic agents in vivo, there were no significant differences between the strains in the tracheal responses to carbachol (P = 0.78) or KCl (P = 0.13) in vitro. Both strains showed inhibition of the isoproterenol relaxation by carbachol (P less than 0.0001). Multiple linear regression analysis showed that the strain that was more sensitive to carbachol in vivo was also more sensitive to isoproterenol in vitro after carbachol contraction (P = 0.014). The greater isoproterenol sensitivity of the tracheae from this strain was not present after contraction with KCl, nor were these tracheae more sensitive to relaxation with sodium nitroprusside. Indomethacin pretreatment of the tissues in vitro augmented the maximal response and the sensitivity to carbachol (P less than 0.001) and KCl (P = 0.0006), and this effect was similar in both strains. Evaluation of isoproterenol relaxation after indomethacin pretreatment was confounded by the lower concentrations of carbachol needed for contraction.(ABSTRACT TRUNCATED AT 250 WORDS)     

Establishment of a multi-dose study of chondroitin sulfate iron colloid for evaluation of the reticuloendothelial system function.

The assay system of chondroitin sulfate iron colloid (CSFe) was established to evaluate the reticuloendothelial system (RES) function in individual rabbits. In the multi-dose study of CSFe, CSFe was repeatedly administered to each individual rabbit with increasing doses (0.6, 1.2, 2.4, 6.0 mg/kg) at set intervals. Blood samples were serially collected after injection of CSFe and the concentration of CSFe in serum was directly measured as an iron concentration by modifying the previously described assay method [1] to minimize the sample volume. The clearance rate of CSFe at each injected dose was computed by the least-squares method and the double-reciprocal plotting of the doses against the phagocytic velocities by the Lineweaver-Burk method was obtained in each rabbit. The maximum phagocytic velocity (Vmax) and the CSFe concentration producing 1/2 Vmax (Kp) obtained in ten rabbits were 0.129 +/- 0.025 mg/kg per min and 0.417 +/- 0.121 mg/kg (mean +/- S.D.), respectively. The results obtained from this multi-dose study were comparable to our previous results obtained from the mean values of five groups given different doses [1]. The clearance rates of CSFe (0.6, 1.2, 6 mg/kg) decreased after the co-injection of 80 mg/kg of carbon colloid. The calculated Vmax and Kp in 29 rabbits were 0.125 mg/kg per min and 1.167 mg/kg. The Kp was apparently greater than that of the control (Vmax = 0.128 mg/kg per min, Kp = 0.421 mg/kg). Carbon colloid (80 mg/kg) was injected to six rabbits after the completion of the first multi-dose study of CSFe and then the second multi-dose study of CSFe was repeated after 24 h. No differences were found in Vmax and Kp between the two studies as were in the control group (10 rabbits) where saline was injected instead of carbon colloid. These results indicated that carbon colloid (80 mg/kg) gives a competitive and reversible inhibition on the RES. This multi-dose study of CSFe may be applicable for a bed-side analysis of the RES function in a patient.     

Studies on the loss of sensitivity in smooth muscle.

Specificity of desensitization effect was investigated at guinea pig ileum using the desensitizing agonists acetylcholine, carbachol, histamine, and serotonine. The desensitization with acetylcholine markedly diminished the contraction effects to test doses of all tested agonists. Contrary to acetylcholine, the histamine desensitization is mainly specific, the serotonine desensitization is a complete specific effect. A different influence of the tested agonists on intracellular ions is discussed.     

Biphasic dose-response relationship for acetylcholine in the chick jugular vein

The pharmacological mechanism of biphasic dose-response relationship for acetylcholine (ACh), relaxation at low doses (1 nM to 0.3 μM) and contraction at high doses (1 μM to 30 μM), in the chick jugular vein was investigated. Neither relaxations nor contractions were affected by the treatment with tetrodotoxin, hexamethonium, d-tubocurarine, phentolamine, propranolol, reserpine, or ouabain. Besides, anoxia did not affect the biphasic pattern of dose-response curve. The contraction was attenuated by the treatment with aspirin or indomethacin, but only slightly. The dose-response curves for these responses to acetylcholine were shifted to the right by the treatment with atropine. Methacholine, carbachol, bethanechol, and arecoline caused similar biphasic responses, although contractions caused by highest doses of bethanechol or arecoline were very small in amplitude. On the other hand, pilocarpine and McN-A-343 only relaxed the strips. The dose-response curves for cholinomimetics were all shifted to the right by the treatment with atropine. It was demonstrated that the responses of the chick jugular vein to muscarinic agonists are different from those of mammalian veins. The mechanisms underlying the biphasic response are discussed.     

Effects of garlic (Allium sativum) extract on the heart rate, rhythm and force of contraction in frog: a dose-dependent study

Garlic juice (dose equivalent to 3.3 g to 33 g garlic) mainly caused bradycardia in frog Rana tigerina. The disturbance in ventricular rhythm was observed prior to than that of atria. Rhythm was specially disturbed at higher doses causing bizarre pattern. Force of contraction of the heart also decreased with higher dose of the garlic extract. The results suggest that garlic extract has some beneficial effect on heart rate modulating the rate, rhythm and force of contraction positively but very high doses may exert non-desirable effects as well.     

Effects of mepacrine on uterine contractile responses

Mepacrine is a potent inhibitor of uterine contractile responses in vitro. Pretreatment of isolated rat uterine horns with mepacrine (1.3 X 10(-4)M) for periods of time ranging from 15 s to 5 min prior to the addition of carbachol (1.0 X 10(-4)M) showed that mepacrine could significantly reduce carbachol-induced uterine contractile responses within 15 s of exposure. The maximal inhibitory effects of mepacrine on uterine contractile responses were observed within 2 min of mepacrine treatment. A dose-response study related to the effect of increasing concentrations of mepacrine (7.5 X 10(-6) to 1.3 X 10(-4)M) on carbachol-induced (1 X 10(-4)M) uterine contractions revealed that a dose of 3.1 X 10(-5)M mepacrine reduced the carbachol-induced contraction by 50%. A dose of 7.8 X 10(-5)M mepacrine produced the maximal inhibitory effect on the carbachol-induced uterine contractions. Two doses of mepacrine (3.1 X 10(-5) and 1.3 X 10(-4)M) significantly reduced maximal contractile responses and shifted contractile dose-response curves of carbachol, oxytocin, prostaglandin F2 alpha, and BaCl2 to the right. Based on the nonselective inhibition by mepacrine of contractile responses induced by different uterotonic agents, these results suggest that mepacrine cannot be used to characterize the role of phospholipase in regulating the actions of hormones in uterine tissue.     

Thermic response of selective muscarinic agonists and antagonists in rat

Effect of some selective agonists and antagonists of cholinergic M receptor subtypes on rectal temperature was investigated in rats at an ambient temperature of 25 degrees +/- 2 degrees C. Centrally administered acetylcholine (ACh) induced transient hypothermia, whereas the muscarinic M1 receptor agonists, arecholine (ip) and McN-A-343 (McN) (icv), induced sustained and dose-related hypothermia. However, the nonspecific muscarinic receptor agonist, oxotremorine, and physostigmine, induced hypothermia at a lower dose and hyperthermia, accompanied by tremors, at higher doses. The muscarinic M2 receptor agonist, carbachol (icv) also produced a dose-related dual effect, hyperthermia and hypothermia being induced by the lower and higher doses, respectively. The M1 receptor antagonists, scopolamine (ip) and pirenzepine (icv), induced hyperthermia, whereas the M2 receptor antagonists, gallamine (icv) and AF-DX 116 (AFDX) (ip), produced hypothermia. The hypothermic effects of ACh. arecholine, McN, physostigmine, oxotremorine and carbachol were attenuated by scopolamine and pirenzepine. However, although scopolamine also inhibited the hyperthermic and tremorogenic effects of the higher dose of oxotremorine, it had a synergistic effect with the hyperthermia-inducing higher dose of physostigmine. AFDX attenuated the hyperthermic effect of the lower dose of carbachol, indicating that it was M2 receptor-mediated. Hemicholinium, an ACh synthesis inhibitor, had a transient hypothermic effect followed by slight hyperthermia. However, it markedly antagonized the hypothermic effects of gallamine and AFDX, indicating that their effects were dependent upon the availability of neuronal ACh. The results indicate that cholinergic hypothermia is a function of central muscarinic M1 receptors, with the M2 receptors serving as automodulators.(ABSTRACT TRUNCATED AT 250 WORDS)     

Stress–strain analysis of jejunal contractility in response to flow and ramp distension in type 2 diabetic GK rats: Effect of carbachol stimulation

Investigation of intestinal motility in a genetic model of GK rats abandons the possible neurotoxic effect of streptozotocin in streptozotocin-induced diabetic model. Seven GK male rats (GK group) and nine normal Wistar rats (Normal group) were used in the study. The motility experiments were carried out in an organ bath containing physiological Krebs solution. Before and after 10⁻⁵ M carbachol application, the pressure and diameter changes of jejunum were obtained in relation to (1) basic contraction, (2) flow-induced contraction with different outlet resistance pressures and (3) contractions induced by ramp distension. The frequency and amplitude of contractions were analyzed from pressure–diameter curves. Distension-induced contraction thresholds and maximum contraction amplitude of basic and flow-induced contractions were calculated in terms of stress and strain. (1) The contraction amplitude increased to the peak value in less than 10 s after adding carbachol. More than two peaks were observed in the GK group. (2) Carbachol decreased the pressure and stress threshold and Young's modulus in the GK group (P<0.01). (3) Carbachol increased the maximum pressure and stress of flow-induced contractions at most outlet pressure levels in both two groups (P<0.001). Furthermore, the flow-induced contractions were significantly bigger at low outlet pressure levels in GK group (P<0.05 and P<0.01). (4) The contraction frequency, the strain threshold and the maximum contraction strain did not differ between the two groups (P>0.05) and between before and after carbachol application (P>0.05). In GK diabetic rats, the jejunal contractility was hypersensitive to flow and distension stimulation after carbachol application.     

A physiological pharmacokinetic model describing the disposition of lycopene in healthy men

A physiological pharmacokinetic model was developed to describe the disposition of lycopene, delivered as a tomato beverage formulation in five graded doses (10, 30, 60, 90, or 120 mg), for a phase I study in healthy male subjects (five per dose). Blood was collected before dose administration (0 h) and at scheduled intervals until 672 h. Serum concentrations of carotenoids and vitamins were measured by high performance liquid chromatography analysis. The model was comprised of seven compartments: gastrointestinal tract, enterocytes, chylomicrons, plasma lipoproteins, fast-turnover liver, slow-turnover tissues, and a delay compartment before the enterocytes. As predicted, the percent absorption at the 10 mg dose (33.9 +/- 8.1%) was significantly greater than at the higher doses; however, the amount of lycopene absorbed (mg) was not statistically different (mean: 4.69 +/- 0.55 mg) between doses, suggesting a possible saturation of absorptive mechanisms. The slow-turnover tissue compartment served as a slow-depleting reservoir for lycopene, and the liver represented the fast-turnover pool. Independent of dose, 80% of the subjects absorbed less than 6 mg of lycopene. This may have important implications for planning clinical trials with pharmacological doses of lycopene in cancer control and prevention if absorption saturation occurs at levels that are already being consumed in the population.     

Evaluation of the reticuloendothelial system function by the vascular clearance of chondroitin sulfate iron colloid

Vascular clearance of chondroitin sulfate iron colloid (CSFe) was evaluated as a test for the reticuloendothelial system (RES) in rabbits. Injected CS59Fe was taken up by the RES in the liver (49%) and bone marrow (41%) after 60 min, suggesting its application for the RES function test. The clearance rate (K-value) of CSFe from the blood was calculated by measuring serum Fe concentrations after releasing iron from CSFe at certain intervals after injection. Depending upon different injected doses, K-values were varied and the phagocytic velocity, computed by multiplying K-values by corresponding injected doses, reached a plateau at high doses, indicating the phagocytosis of CSFe by the RES takes a saturation process. Double-reciprocal plotting of the dose and the phagocytic velocity showed a linear relationship, which provided the data on the maximum phagocytic velocity (Vmax), 0.122 mg/kg/min, and the CSFe concentration producing 1/2 Vmax (Kp), 0.426 mg/kg. Thus, this CSFe clearance test can be used for the evaluation of the RES function.     

Endothelin, PAF and thromboxane A2 in allergic pulmonary hyperreactivity in mice

The role of endothelin, PAF and thromboxane A2 in airway hyperreactivity (AHR) to carbachol induced by ovalbumin sensitization and challenge in Balb/c mice was investigated. Ovalbumin sensitization and challenge induced significant AHR to carbachol in actively sensitized and challenged mice. Treatment of these mice with the PAF antagonist CV-3988 (10 microg kg(-1), i.v.) completely abolished OVA-induced AHR to carbachol. Treatment of sensitized mice with the TxA2 antagonist L-654,664 (1 mg kg(-1), i.v.) partially blocked the induction of AHR in OVA-challenged mice. The intranasal administration of 50 pmol of the ET(A) receptor antagonist BQ-123 had no effect on the PIP but produced a significant reduction at the dose of 100 pmol. The intravenous administration of BQ-123 (100 pmol) reduced the PIP only at the highest doses of carbachol. The ET(B) receptor antagonist BQ-788 administered either via the intranasal or intravenous route had no effect on the PIP at the dose of 100 pmol. Na?ve mice treated with either U-44069 (25 or 100 microg kg(-1), i.v.), endothelin-1 (100 pmol, intranasally) or the ET(B) receptor agonist IRL-1620 (100 pmol, intranasally) showed a marked increase in airway reactivity to carbachol. These results suggest an important role for endothelin, PAF and thromboxane A2 in AHR in mice actively sensitized and challenged with ovalbumin.     

Instantaneous reactions of guinea-pig vas deferens preparation to X-irradiation

Summary In the mechanical-inactive guinea-pig vas deferens X-rays (25 kV) at threshold doses of about 100 kR initiated phasic activity and an immediate increase in tone. After addition of acetylcholine, noradrenaline or adrenaline to the rinsing solution a slight contractile activity of vas deferens appeared and the preparation reacted to X-irradiation at a threshold dose of 3 to 5 kR (dose-rate 20 kR/min) with increased phasic contractions and with a dose and dose-rate dependent tonic contraction. After repeated irradiation a sensitization was observed. X-rays produced tonic contractions of the vas deferens preparation up to a total dose of about 200 kR (fractionated irradiation). An irreversible contraction of the vas appeared after continuous exposure to X-ray doses larger than 500 kR (dose-rate 20 kR/min).     

TRH dissociates the aggressivity of vegetative and motor phenomena induced by carbachol in the cat

In these experiments interaction of thyrotropin releasing hormone (TRH) and carbachol injected into the cerebral ventricles of unanaesthetized cats has been investigated. Intracerebroventricular (i.c.v.) carbachol as well as i.c.v. TRH produced emotional behaviour, autonomic and motor phenomena. The most impressive feature of i.c.v. carbachol was the aggressive behaviour, whereas that of i.c.v. TRH the autonomic changes. In cats treated with i.c.v. TRH, the aggressive behaviour, but the autonomic and motor changes of i.c.v. carbachol was potentiated. Since there is evidence that carbachol acts mainly on muscarinic M-2 receptors, the potentiation by TRH of aggressive behaviour, but not the autonomic and motor changes induced by carbachol could indicate heterogeneity of central muscarinic M-2 receptors.     

Toxicology of the prostaglandins

Available published material of adverse reactions to prostaglandins (PGs) at various dosages routes and levels is reviewed. Animal studies are of 2 kinds: studies of pharmacological effects and studies of toxicological reactions (i.e., acute dose levels). The pharmacology of PGs show the drugs have 3 areas of action: 1) smooth muscle effects, either contraction or relaxation (cardiovascular effects and reproductive tract; relaxation of vascular smooth muscles by some PGs and contraction by others); 2) nervous system effects; and 3) lipid and carbohydrate metabolism. In humans, PGE1 was administered intravenously and it was found that adverse effects were related to rate of administration rather than to total dose; side effects included flushing, headache, visual disturbances, and restlessness, factors which might suggest a correlation with central nervous system effects (as found in animal studies). PGE1 administered at acute doses orally, by inhalation, and intradermally show effects attributable to gastrointestinal disturbances, respiratory problems, and erythematous responses. In general, studies in humans of other PGs have found similar adverse reactions, all of which are explained by known mechanisms of action of PGs. Dose levels constituting acutity are dependent on route (i.e., oral doses are much higher than intravenous doses), with rapid intravenous infusion causing the most adverse reactions.     

Involvement of phosphorylation of myosin phosphatase by ROCK in trabecular meshwork and ciliary muscle contraction

The control of smooth muscle contraction is an important factor in maintaining normal intraocular pressure. However, the specific factors causing changes in control by phosphorylation/dephosphorylation schemes in the eye are not well-defined. The purposes of this experiment were to (i) determine the localization of ROCK (Rho-associated, coiled coil-forming kinase) in monkey and rabbit eye tissues and (ii) measure phosphorylation of ROCK substrate during trabecular meshwork or ciliary muscle contraction induced by carbachol. We found that mRNAs for both ROCK I and II were expressed in most eye tissues from rabbit and monkey. Proteins for ROCK I and II were present in all eye tissues studied except lens. When trabecular meshwork or ciliary muscle were incubated with carbachol to induce contraction, phosphorylation of the myosin-binding subunit (MBS) of myosin phosphatase, a substrate for ROCK, started within 1 min and continued for at least 1 h. This phosphorylation was well correlated with contraction of trabecular meshwork or ciliary muscle. These results suggested that ROCK might regulate contraction of trabecular meshwork or ciliary muscle through phosphorylation of MBS of myosin phosphatase.