Synthesis and pharmacological activity of a series of dihydro-1H-pyrrolo[1,2-a]imidazole-2,5(3H,6H)-diones, a novel class of potent cognition enhancers.

A series of dihydro-1H-pyrrolo[1,2-a]imidazole-2,5(3H,6H)-diones were synthesized. These bicylic derivatives contain both the 2-pyrrolidinone and 4-imidazolidinone nuclei, already recognized as important for cognition enhancing activity. In addition, these structures maintain the backbone of piracetam and oxiracetam with the acetamide side chain restricted in a folded conformation. Their ability to reverse scopolamine-induced amnesia was assessed in a one trial, step-through, passive avoidance paradigm. The main features observed are a potent antiamnestic activity after ip administration (minimal effective dose being between 0.3 and 1 mg/kg ip for most compounds), the presence of a bell-shaped dose-response curve and, generally, a reduction of biological activity after po administration. However, the unsubstituted compound (15, dimiracetam) shows no evidence of a bell-shaped dose-response curve and completely retains activity when given orally, being 10-30 times more potent than the reference drug oxiracetam.

Oxiracetam antagonizes the disruptive effects of scopolamine on memory in the radial maze.

The effects exerted by oxiracetam on the disruption of performance induced by scopolamine in the radial arm maze were investigated in overtrained rats. Scopolamine induced a dose-related decrease in the efficiency of responding and an increase of running time. The effect of the SC injection of 0.2 mg/kg scopolamine on the efficiency of responding was antagonized by the IP administration of 30 mg/kg oxiracetam, while the effect on running time induced by the same dose of scopolamine was not. Physostigmine (0.3 mg/kg SC) antagonized both effects of 0.2 mg/kg scopolamine. Methylscopolamine, at the dose of 0.2 mg/kg SC, was devoid of any effect on both parameters. Increasing the dose of methylscopolamine to 0.63 mg/kg did cause serious peripheral effects which eventually prevented some animals from completing the task. Similar peripheral effects were observed after administration of 0.63 mg/kg scopolamine. The effects of this dose of scopolamine on efficiency and running time were not antagonized by pretreatment with 100 mg/kg oxiracetam. Oxiracetam alone (30 or 100 mg/kg IP) did not modify the performance of previously trained rats. The present results suggest that oxiracetam selectively restores cholinergic mechanisms which are involved in learning and memory.

Brain entry and direct central pharmacological effects of the nootropic drug oxiracetam. Oxiracetam: brain entry and pharmacological effects.

Oxiracetam administered systemically to rats as a 14C radiolabelled drug (200 mg/kg p.o. or 100 mg/kg intra-artery) enters the brain and is found unmetabolized above all in some of the brain areas functionally involved in the modulation of cognitive processes. Among the brain areas examined, the largest amount of compound was recovered in septum, followed by hippocampus; a smaller amount was found in cerebral cortex and striatum. 14C oxiracetam administered directly into the lateral ventricles of the brain presented a similar pattern of distribution, indicating that the tropism of the drug for the above brain areas does not depend on its route of administration. The amounts of oxiracetam that could reach the brain after systemically administered pharmacologically active doses were also estimated. These amounts (1.9 to 19 nmols/rat) were delivered through a permanently implanted cannula, into the lateral ventricles of the brains of conscious, freely moving rats. In this experimental condition oxiracetam dose-dependently antagonized the amnesia induced by scopolamine (0.66 mg/kg s.c.). The above findings clearly indicate that oxiracetam enters the brain and directly exerts its pharmacological activity there.

The effect of oxiracetam treatment on alterations of lipid metabolism in brain areas from spontaneously hypertensive rats.

It has been previously demonstrated that spontaneously hypertensive adult rats (SHR) develop severe hypertension and cerebrovascular lesions on drinking 1% NaCl from weaning and that the phospholipid metabolism in the whole brain is actively altered in these lesioned animals (SHR-NaCl) as compared to SHRs which drink only water and show only sporadic cerebrovascular lesions. We have now assayed the incorporation of labelled choline, ethanolamine, glycerol and arachidonic acid into the phospholipids from the cortex and hippocampus of SHR-water and SHR-NaCl at different time intervals from injection into the lateral ventricle of the brain. A noticeable decrease of both choline and arachidonate specific activity (SA) in the phospholipids was found in the cortex and hippocampus (where the effect is most evident) from SHR-NaCl. Based on the literature and the data obtained, we suggest that in SHR-NaCl brain areas a release of choline and fatty acid also occurs from choline glycerophospholipids as a consequence of the cerebrovascular lesions caused by NaCl treatment. Even if a relatively minor loss of the amount of the lipids studied is evident from our results as compared to their entire pool, this change may be quite important if it causes a modification of the lipidic bilayer in excitable membranes. In a parallel group of SHR-NaCl animals, treated with the nootropic drug oxiracetam, we observed that the metabolic utilization of the precursors was completely restored. These experimental data favour the hypothesis that oxiracetam is effective in stimulating the phospholipid metabolism rate at levels even higher than those of the SHR-water animals.

Renal function studies with cadralazine in conscious dogs: a comparison with hydralazine.

The effect of the antihypertensive vasodilator, cadralazine, on renal function in the conscious dog was compared to that of hydralazine using inulin and para-aminohippurate clearances. Both drugs were administered as intravenous bolus, at the dose of 1 mg/kg. As expected, hydralazine rapidly decreased mean blood pressure (from 110 to 89 mmHg), significantly increased heart rate (from 109 to 190 beats/min), markedly decreased urine volume (from 0.90 to 0.47 ml/min) and sodium excretion (from 101 to 45 microEq/min), and increased potassium excretion (from 28 to 53 microEq/min). Cadralazine displayed a similar activity on blood pressure and on heart rate, but differently from hydralazine, these effects appeared more slowly and were not accompanied by sodium and water retention. Hydralazine, but not cadralazine, caused a quick and transient decrease in glomerular filtration rate (from 55 to 40 ml/min), whereas both compounds increased renal plasma flow and reduced renal vascular resistance, renal extraction of para-aminohippurate and filtration fraction. Moreover, cadralazine increased plasma renin activity to a lesser extent than hydralazine, and this could explain the different effect on water and sodium excretion after acute administration of the two drugs.

Effect of age and strain on cerebral phospholipid metabolism.

It has been demonstrated that spontaneously hypertensive adult rats (SHR) develop severe hypertension and cerebrovascular lesions on drinking 1% NaCl from weaning. Phospholipid metabolism is actively altered in these severely lesioned animals (SHR-NaCl) as compared to SHRs which drink only water and showed only sporadic cerebrovascular lesions. We have tested the incorporation of water soluble phospholipid precursors into the corresponding phospholipid from different brain areas, by injecting either a mixture of labeled glycerol and choline or glycerol and ethanolamine into the lateral ventricle of the brain of adult (4 months old) and senescent (12 months old) SHR-NaCl. The results were compared to those obtained from 4 and 12 months old Wistar normotensive rats. When adult normotensive rats were compared with adult hypertensive rats (4-SHR-NaCl) incorporation was found to decrease in some areas according to the precursors injected. Similar results were obtained from 12 month old normotensive Wistar rats that, however, showed a decrease in phospholipid biosynthesis in all the area tested. Interestingly, no significant differences of incorporation rate were found between 12 month old normotensive and 12 month old hypertensive rats.

Experimental behavioral studies with oxiracetam on different types of chronic cerebral impairment.

Oxiracetam (4-hydroxy-2-oxo-1-pyrrolidine acetamide) is a novel compound effective in improving learning and memory in normal animals as well as in animals with acute cerebral impairment induced by a variety of noxious stimuli (i.e., electroshock, neurodepressants, metabolic inhibitors, hypoxia). In accordance with a stepwise approach to the pharmacological studies, the compound was also tested in rats with chronic cerebral impairment due to aging, cerebrovascular lesions, and congenital microencephaly. To evaluate the brain telencephalic functions, the learning rate of two conditioned avoidance responses using the pole climbing test and the performance in a multiple-choice water maze were considered. Oxiracetam at doses ranging from 10 to 60 mg/kg i.p. improved the learning rate of these animals with impaired cognitive functions. The compound was active also by oral administration.

Methylazoxymethanol microencephaly in rats: neurochemical characterization and behavioral studies with the nootropic oxiracetam.

The administration of methylazoxymethanol (MAM) to pregnant rats induced a marked reduction in the weight of the offspring's brain. This reduction was due to aplasia of the cortex and hippocampus, whose thicknesses were 50% of those of control animals. A significant reduction was also observed in the striatum. This aplasia could be ascribed to the antimitotic effect of MAM, which, when given at gestational day 15, prevented the development of neurons in the three brain areas mentioned. Indeed, we infer here that the total number of GABA-receptor complexes, as measured by [3H]muscimol and [3H]flunitrazepam binding, was reduced to the same degree as was the weight of the cortex. Similarly, total [3H]haloperidol binding sites were reduced in the striatum. From the behavioral point of view, offspring of MAM-treated rats (MAM rats) showed impaired acquisition in the water-maze and pole-climbing tests, indicating that this brain aplasia had disrupted cognitive processes. In contrast, these animals showed normal growth, and grossly their behavior appeared normal. Oxiracetam, a new compound that belongs to the recently described class of nootropic drugs, was able to restore acquisition processes in MAM rats. We propose therefore that MAM rats might become an interesting and quite simple animal model for evaluation of new acquisition-enhancing drugs. Moreover, this model could also be useful to study the neurochemical correlates of cognitive processes.

Effects of pildralazine alone and in combination on severe hypertension and cerebrovascular lesions in saline-drinking spontaneously hypertensive rats.

The efficacy of long-term treatments with the vasodilator antihypertensive pildralazine alone and in combination with propranolol or dihydrochlorothiazide in preventing blood pressure increase and cerebrovascular lesions in saline-drinking spontaneously hypertensive rats (SHR) was evaluated. Groups of 20-40 animals were treated daily for 13 weeks. At the end of the experiments the brains were examined histologically and vascular lesions, ranging from arterial wall damage to cerebral infarction, were checked. In the control groups (n = 60), systolic blood pressure gradually rose to over 200 mmHg, and about 74% of the animals had lesions. Pildralazine at a dose of 1 mg/kg p.o. significantly inhibited the onset of severe hypertension; the combination of pildralazine with moderately effective doses of propranolol (10 mg/kg p.o.) or dihydrochlorothiazide (5 mg/kg p.o.) completely prevented the blood pressure increase. When blood pressure control was achieved, the incidence of cerebrovascular lesions was also significantly reduced.

Hemodynamic profile of the antihypertensive vasodilator cadralazine in conscious dogs.

Hemodynamic activity of cadralazine (ethyl-2-[6-(2-hydroxypropyl)-ethylaminol-3-pyridazinyl hydrazine carboxylate), a new, long-acting antihypertensive agent, was evaluated on systemic and regional circulation in conscious dogs. Cadralazine given i.v. (1 mg/kg) caused a sustained fall in total peripheral vascular resistances and mean blood pressure (from 103 to 90 mmHg) and an increase in heart rate (from 97 to 161 beats/min). Heart rate variations paralleled the drop in peripheral resistances. Cadralazine produced a consistent increase in cardiac output, and this effect was related to the increase in heart rate. No significant change in myocardial contractility was observed. Blood flow was increased and vascular resistances decreased in coronary, iliac and mostly in renal vascular beds, whereas the variations in the mesenteric district were not significant. This hemodynamic pattern characterizes cadralazine as a vasodilator. Changes in hemodynamic responses to epinephrine (1 microgram/kg i.v.) after cadralazine treatment were also evaluated. Cadralazine reduced hypertension and bradycardia effects, increased hypotension and tachycardia responses, and caused a further increase in cardiac output and coronary blood flow. These effects of cadralazine are not due to alpha-blocking or to beta-stimulating properties.

Cyclic GABA-GABOB analogues. IV. Activity on learning and memory.

A number of N-substituted 4-hydroxy-, 4-acyloxy- and 4-alkoxy-2-pyrrolidinones were examined by a screening method predictive of their activity on cognitive processes. The 1-aminocarbonylmethyl-substituted compounds showed a favorable effect on learning and memory, and among them the most active was the 4-hydroxy derivative, oxiracetam, which had a potency considerably higher than piracetam, used for comparison purposes.

A screening method for substances potentially active on learning and memory.

A series of substances from different pharmacological classes was examined by two different tests for their activity on learning and memory. A concomitant evaluation was performed by use of a one-trial passive avoidance immediately followed by electroshock in mice and the pole-climbing test in rats. Only doses not producing changes in gross behavior (Irwin's test) were used. A facilitating action on all the considered parameters was observed for d-amphetamine, caffeine, L-glutamine, Mg pemoline, phosphorylserine, piracetam, strychnine, and tricyanoaminopropene. A worsening action was found for atropine, cycloheximide, diazepam, and morphine. Chlorpheniramine, diphenylhydantoin, GABA, imipramine, meclizine, mescaline, metrazol, and testosterone showed no or doubtful activity. Our results suggest that the parallel use of these relatively simple tests supplies information that is satisfactory for screening purposes and that has a high degree of predictability as substantiated by literature data.

[Effect of S-carboxymethylcysteine on blood levels of amoxycillin and its diffusion through lung parenchyma and pleural exudate in the rat]. / Influenza della S-carbossimetilcisteina sui livelli ematici di amoxicillina e sulla sua diffusibilità nel parenchyma polmonare ed essudato pleurico nel ratto.

The fixed combination (10 : 3) of amoxycillin and S-carboxymethylcysteine, which is indicated in the therapy of the respiratory tract infections, was given to rats by oral route to determine the blood levels and the concentrations in the pulmonary parenchyma and pleural exudate. In comparison with the antibiotic alone the combination determined higher levels of amoxycillin in the blood (0.5, 1.2 hours after administration) in the homogenized lung tissue (2 hours after administration) and in pleural inflammatory exudate (0.5, 1, 2 hours after administration) caused by intrapleural injection of carrageenan. The enhanced distribution of the antibiotic in the target tissues further supports the clinical usefulness of this combination.

Pharmacological studies on cadralazine: a new antihypertensive vasodilator drug.

Cadralazine is a new, orally effective antihypertensive vasodilator. Acute experiments indicate that the compound reduces blood pressure and increases heart rate. The doses which reduce systolic blood pressure by 25% (ED25) are very similar after oral and intravenous administration (spontaneously hypertensive rats, 1.8 mg/kg, p.o.; 2.3 mg/kg, i.v.; renal hypertensive dogs, 0.26 mg/kg, p.o.; 0.24 mg/kg, i.v.; awake normotensive dogs, 0.98 mg/kg, p.o.; 1.01 mg/kg, i.v.). By both routes the peak effect is reached after 3-5 hr, and the activity lasts more than 24 hr. Repeated oral administration in spontaneously hypertensive rats reduces blood pressure with no evidence of tolerance. Cadralazine reverses hypertensive responses to epinephrine in awake normotensive dogs and anesthetized cats. The inhibition of the increase in blood pressure induced by sympathetic outflow activation in pithed rats parallels the antihypertensive activity in onset, intensity, and duration. The effects of cadralazine are not due to a blockade of alpha-adrenoceptors, sympathetic neurons, or ganglionic transmission. Cadralazine has no antihistaminic, anticholinergic, or spasmolytic activity and no specific effect on behavioral tests. In comparison with hydralazine, cadralazine has less acute toxicity and a greater activity by the oral route.

Renal function studies with propildazine alone and in combination with propranolol in dogs.

The vasodilator antihypertensive propildazine acutely administered in awake dogs (0.1 mg/kg i.v.) produced an arterial blood pressure drop accompanied by tachycardia, decreased urine volume and urinary sodium excretion, and increased urinary potassium excretion. A transient, quickly reversible decrease in glomerular filtration rate was observed; on the contrary, the reduction in urine volume and urinary sodium excretion was longer lasting. There was indirect evidence that sodium retention was mainly caused by increased tubular reabsorption. Decreases in renal resistance, renal extraction of p-aminohippurate, and filtration fraction were observed together with an increase in renal plasma flow. Propildazine at the above dose in combination with propranolol (1 mg/kg i.v.) had a hypotensive effect with a smaller increase in heart rate and without significant variations in urine volume or urinary sodium excretion. These effects could be attributed to antagonism of the reflex-activated sympathoadrenal system by propranolol. In these conditions, the glomerular filtration rate was not significantly modified, while the effects on other renal parameters were similar to those observed with propildazine.

[3-Hydrazinepyridazine derivatives. III. Synthesis and anti-hypertensive activity of new 3-(2-acylhydrazine)-pyridazine-6-alkylamino substitutes]. / Derivati della 3-idrazinopiridazina. Nota III. Sintesi ed attività antiipertensiva di nuove 3-(2-acilidrazino)-piridazine-6-alchilamino sostituite.

The synthesis of 2-(6-dialkylamino-3-pyridazinyl)hydrazinecarboxylates (II) and hydrazides (V) from the corresponding 3-chloro-6-dialkylaminopyridazines (I) is described. The 6-substituted derivatives of 2,3-dihydro-1,2,4-triazolo[4,3-b]pyridazine-3-ones (III) and 1,2,4-triazolo[4,3-b)pyridazines (VI) were obtained by thermal cyclization of (II) and (V), respectively. The new acyl derivatives were evaluated together with todralazine and budralazine as antihypertensive agents in comparison with propildazine and hydralazine. The ethoxycarbonyl compound [(II g), ISF 2469] exhibits good antihypertensive activity particularly via oral administration. Its interesting pharmacodynamic properties, including slow onset and long-lasting action, qualify it for further pharmacological and clinical studies.

[Derivatives of 3-hydrazinopyridazine. II. Synthesis and antihypertensive activity of new 3-hydrazino-6-monoalkylaminopyridazines]. / Derivati della 3-idrazinopiridazina. Nota II - Sintesi e attività antiipertensiva di nuove 3-idrazino-6-monoalchilaminopiridazine.

The synthesis of some 3-hydrazino-6-monoalkylaminopyridazines (VIII) by reaction of 3,6-dichloropyridazine with various N-monoalkylbenzylamines, followed by reaction with hydrazine and final debenzylation in an acidic medium, is described. The new compounds were evaluated as antihypertensive agents and exhibited the same or higher activity than hydralazine, but lower activity than propildazine (ISF 2123).