Antinociceptive Effect of the Combination of a Novel α4ß2* Agonist with Donepezil in a Chronic Pain Model.

Chronic pain presents a major challenge in contemporary medicine, given the limited effectiveness and numerous adverse effects linked to available treatments. Recognizing the potential of the cholinergic pathway as a therapeutic target, the present work evaluates the antinociceptive activity of a combination of Cris-104, a novel α4ß2* receptor agonist, and donepezil, a central anticholinesterase agent. Isobolographic analysis revealed that equimolar combination was approximately 10 times more potent than theoretically calculated equipotent additive dose. Administration of Cris-104 and donepezil combination (3 µmol/kg) successfully reversed hyperalgesia and mechanical allodynia observed in rats subjected to spinal nerve ligation (SNL). The combination also modulated neuroinflammation by reducing astrocyte activation, evident in the decreased expression of glial fibrillary acidic protein (GFAP) in the spinal cord. The observed synergism in combining a nicotinic receptor agonist with an anticholinesterase agent underscores its potential for treating chronic pain. This alternative therapeutic distinct advantage, including dose reduction and high selectivity for the receptor, contribute to a more favorable profile with minimized adverse effects.

Reduction of cardiac and renal dysfunction by new inhibitor of DPP4 in diabetic rats.

BACKGROUND: Increased mortality due to type 2 diabetes mellitus (T2DM) has been associated with renal and/or cardiovascular dysfunction. Dipeptidyl dipeptidase-4 inhibitors (iDPP-4s) may exert cardioprotective effects through their pleiotropic actions via glucagon-like peptide 1-dependent mechanisms. In this study, the pharmacological profile of a new iDPP-4 (LASSBio-2124) was investigated in rats with cardiac and renal dysfunction induced by T2DM. METHODS: T2DM was induced in rats by 2 weeks of a high-fat diet followed by intravenous injection of streptozotocin. Metabolic disturbance and cardiac, vascular, and renal dysfunction were analyzed in the experimental groups. RESULTS: Sitagliptin and LASSBio-2124 administration after T2DM induction reduced elevated glucose levels to 319.8 ±â€¯13.2 and 279.7 ±â€¯17.8 mg/dL, respectively (p < 0.05). LASSBio-2124 also lowered the cholesterol and triglyceride levels from 76.8 ±â€¯8.0 to 42.7 ±â€¯3.2 mg/dL and from 229.7 ±â€¯25.4 to 100.7 ±â€¯17.1 mg/dL, in diabetic rats. Sitagliptin and LASSBio-2124 reversed the reduction of the plasma insulin level. LASSBio-2124 recovered the increased urinary flow in diabetic animals and reduced 24-h proteinuria from 23.7 ±â€¯1.5 to 13.3 ±â€¯2.8 mg (p < 0.05). It also reduced systolic and diastolic left-ventricular dysfunction in hearts from diabetic rats. CONCLUSION: The effects of LASSBio-2124 were superior to those of sitagliptin in the cardiovascular systems of T2DM rats. This new prototype showed promise for the avoidance of comorbidities in a T2DM experimental model, and thus may constitute an innovative therapeutic agent for the treatment of these conditions in the clinical field in future.

Human Mesenchymal Stem Cell Therapy Reverses Su5416/Hypoxia-Induced Pulmonary Arterial Hypertension in Mice.

Aims: Pulmonary arterial hypertension (PAH) is a disease characterized by an increase in pulmonary vascular resistance and right ventricular (RV) failure. We aimed to determine the effects of human mesenchymal stem cell (hMSC) therapy in a SU5416/hypoxia (SuH) mice model of PAH. Methods and Results: C57BL/6 mice (20-25 g) were exposure to 4 weeks of hypoxia combined vascular endothelial growth factor receptor antagonism (20 mg/kg SU5416; weekly s.c. injections; PAH mice). Control mice were housed in room air. Following 2 weeks of SuH exposure, we injected 5 × 105 hMSCs cells suspended in 50 µL of vehicle (0.6 U/mL DNaseI in PBS) through intravenous injection in the caudal vein. PAH mice were treated only with vehicle. Ratio between pulmonary artery acceleration time and RV ejection time (PAAT/RVET), measure by echocardiography, was significantly reduced in the PAH mice, compared with controls, and therapy with hMSCs normalized this. Significant muscularization of the PA was observed in the PAH mice and hMSC reduced the number of fully muscularized vessels. RV free wall thickness was higher in PAH animals than in the controls, and a single injection of hMSCs reversed RV hypertrophy. Levels of markers of exacerbated apoptosis, tissue inflammation and damage, cell proliferation and oxidative stress were significantly greater in both lungs and RV tissues from PAH group, compared to controls. hMSC injection in PAH animals normalized the expression of these molecules which are involved with PAH and RV dysfunction development and the state of chronicity. Conclusion: These results indicate that hMSCs therapy represents a novel strategy for the treatment of PAH in the future.

Novel agonist of α4ß2* neuronal nicotinic receptor with antinociceptive efficacy in rodent models of acute and chronic pain.

OBJECTIVE: To demonstrate the antinociceptive and antihypersensitivity mechanisms of Cris-104 (1-{2-[5-(4-fluorophenyl)-1H-pyrazol-4-yl]ethyl}piperidine), a novel selective α4ß2* nicotinic acetylcholine receptor (nAChR) agonist, in rodent acute/inflammatory and chronic pain models. MATERIALS AND METHODS: Hot-plate and formalin tests in mice were used to examine Cris-104-induced antinociceptive effects on thermal/inflammatory pain. Cris-104 effects on hypersensitivity, norepinephrine (NE) release in the spinal dorsal horn, and neuronal activity in the locus coeruleus (LC) were examined in rats with lumbar spinal nerve ligation using behavioral, microdialysis, and extracellular recording methods. Cris-104 effects on spontaneous locomotion were examined in an open-field test. RESULTS: Cris-104 induced dose-dependent antinociception effects in hot-plate and formalin tests, and these effects were blocked by the general nAChR antagonist mecamylamine, the selective α4ß2* nAChR antagonist dihydro-beta-erythroidine, and the α2-adrenoceptor antagonist yohimbine, but not by the α1-adrenoceptor antagonist prazosin. Systemic and spinally perfused Cris-104 increased NE concentrations in microdialysates from the spinal cord in both normal and SNL rats. Systemic Cris-104 increased neuronal activity in the LC of normal rats. Mecamylamine blocked the effects of Cris-104 on spinal NE release and LC neuronal activity. Systemic Cris-104 did not affect locomotor activity significantly. CONCLUSION: The α4ß2 neuronal nAChR agonist, Cris-104, was effective for treatment of pain via descending noradrenergic inhibition of pain signaling.

Cardioprotection Induced by Activation of GPER in Ovariectomized Rats With Pulmonary Hypertension.

Pulmonary hypertension (PH) is a disease of women (female-to-male ratio 4:1), and is associated with cardiac and skeletal muscle dysfunction. Herein, the activation of a new estrogen receptor (GPER) by the agonist G1 was evaluated in oophorectomized rats with monocrotaline (MCT)-induced PH. Depletion of estrogen was induced by bilateral oophorectomy (OVX) in Wistar rats. Experimental groups included SHAM or OVX rats that received a single intraperitoneal injection of MCT (60 mg/kg) for PH induction. Animals received s.c. injection of either vehicle or G1, a GPER agonist, (400 µg/kg/day) for 14 days after the onset of disease. Rats with PH exhibited exercise intolerance and cardiopulmonary alterations, including reduced pulmonary artery flow, biventricular remodeling, and left ventricular systolic and diastolic dysfunction. The magnitude of these PH-induced changes was significantly greater in OVX versus SHAM rats. G1 treatment reversed both cardiac and skeletal muscle functional aberrations caused by PH in OVX rats. G1 reversed PH-related cardiopulmonary dysfunction and exercise intolerance in female rats, a finding that may have important implications for the ongoing clinical evaluation of new drugs for the treatment of the disease in females after the loss of endogenous estrogens.

Antihyperalgesic effects of a novel muscarinic agonist (LASSBio-873) in spinal nerve ligation in rats.

New chemicals or adjuvants with analgesic effects on chronic pain are needed and clinically relevant due to the limited number of effective compounds that possess these characteristics. LASSBio-873, a pyrazolo[3,4-b]pyrrolo[3,4-d]pyridine derivative, activates muscarinic cholinergic receptors and has potent analgesic effects on acute and inflammatory pain. The present study evaluated the therapeutic and prophylactic effects of oral administration of LASSBio-873 in a spinal nerve ligation (SNL) model of chronic peripheral nerve injury. LASSBio-873 (100 mg/kg) inhibited the development of thermal hyperalgesia and mechanical allodynia when administered once daily for 7 consecutive days after SNL surgery and reversed these symptoms. LASSBio-873 treatment did not alter rat behaviour in open field testing measured during the first 24 h after administration and again after 7 continuous days administration. The analgesic effect of LASSBio-873 was inhibited by intrathecal methoctramine, an M2 receptor antagonist, implicating the muscarininc M2 receptor signalling pathway in the drug's action. These results reinforce the potential of LASSBio-873 as a possible prototype for the development of more effective alternatives for the treatment of neuropathic pain.

Docking, synthesis and anti-diabetic activity of novel sulfonylhydrazone derivatives designed as PPAR-gamma agonists.

Diabetes is a metabolic disorder characterized by hyperglycemia. When not properly controlled, complications include neuropathy, coronary artery disease, and renal failure. Several drugs are approved for diabetes treatment; however their use is associated with side effects and lack of efficacy in attenuating the development of long-term complications. This work describes the virtual screening and synthesis of a novel series of sulfonylhydrazone derivatives designed as peroxisome proliferator-activated receptor gamma (PPARγ) agonists and investigation of the analogs for hypoglycemic activity in a murine model of diabetes. Docking studies identified LASSBio-331 (5) as having theoretical affinity for PPARγ similar to the prototype (S)-rosiglitazone. Several structural modifications were proposed for the structure of LASSBio-331, resulting in the synthesis of five novel compounds, which showed experimental affinity for PPARγ. Among these new compounds, LASSBio-1471 (15) had the best theoretical binding energy for PPARγ and was selected for testing in STZ-induced diabetes. Four weeks after single intravenous injection of STZ (60 mg/kg), Wistar rats were treated with vehicle (DMSO) or LASSBio-1471 (20 mg/kg, i.p.) for 7 days. The blood glucose levels of rats treated with LASSBio-1471 were reduced from 548.4 ± 26.0 mg/dL before treatment to 259.6 ± 73.1 mg/dL (P < 0.05). Paw withdrawal threshold was significantly reduced in diabetic rats and was restored from 21.9 ± 1.7 g to 36.7 ± 1.2 g after 7 days of treatment with LASSBio-1471 (P < 0.05). Thus, the novel sulfonylhydrazone derivative is a PPARγ ligand that is effective for treatment of diabetic neuropathy in STZ-injected rats.

Inhibition of L-type calcium current by tramadol and enantiomers in cardiac myocytes from rats.

BACKGROUND: Tramadol is a centrally acting analgesic, whose mechanism of action involves opioid-receptor activation. Previously, we have shown that tramadol and its enantiomers had a negative inotropic effect on the papillary muscle in which the (+)-enantiomer is more potent than (-)- and (±)-tramadol. OBJECTIVE: In this study, we investigated the effects of tramadol and its enantiomers on L-type calcium current (ICa-L). METHODS: The experiments were carried out in isolated Wistar rat ventricular myocytes by using the whole cell patch clamp technique. RESULTS: Tramadol (200 µM) reduced the peak amplitude of ICa-L at potentials from 0 to +50 mV. At 0 mV, I(Ca-L) was reduced by 33.7 ± 7.2%. (+)- and (-)-tramadol (200 µM) produced a similar inhibition of ICa-L, in which the peak amplitude was reduced by 64.4 ± 2.8% and 68.9 ± 5.8%, respectively at 0 mV (p > 0.05). Tramadol, (+)- and (-)-tramadol shifted the steady-state inactivation of ICa-L to more negative membrane potentials. Also, tramadol and (+)-tramadol markedly shifted the time-dependent recovery curve of I(Ca-L) to the right and slowed down the recovery of I(Ca-L) from inactivation. The time constant was increased from 175.6 ± 18.6 to 305.0 ± 32.9 ms (p < 0.01) for tramadol and from 248.1 ± 28.1 ms to 359.0 ± 23.8 ms (p < 0.05) for (+)-tramadol. The agonist of µ-opioid receptor DAMGO had no effect on the I(Ca-L). CONCLUSION: The inhibition of ICa-L induced by tramadol and its enantiomers was unrelated to the activation of opioid receptors and could explain, at least in part, their negative cardiac inotropic effect.

Inibição da corrente de cálcio tipo l por tramadol e enantiômeros em miócitos cardíacos de ratos / Inhibition of L-type calcium current by tramadol and enantiomers in cardiac myocytes from rats

FUNDAMENTO: O tramadol é um analgésico de ação central cujo mecanismo de ação envolve a ativação de um receptor opioide. Anteriormente, mostramos que o tramadol e seus enantiômeros apresentavam um efeito inotrópico negativo sobre o músculo papilar no qual o (+)-enantiômero era mais potente que (-)- e (±)-tramadol. OBJETIVO: No presente trabalho, investigamos os efeitos do tramadol e seus enantiômeros na corrente de cálcio tipo L (I Ca-L). MÉTODOS: Os experimentos foram realizados em miócitos ventriculares isolados de ratos Wistar utilizando a técnica de patch-clamp com configuração de célula inteira. RESULTADOS: O tramadol (200 µM) reduziu a amplitude de pico do I Ca-L em potenciais de 0 a +50 mV. Em 0 mV, a I Ca-L foi reduzida em 33,7 ± 7,2 por cento. (+)- e (-)-tramadol (200 µM) produziram uma inibição semelhante da I Ca-L, na qual a amplitude do pico foi reduzida em 64,4 ± 2,8 por cento e 68,9 ± 5,8 por cento, respectivamente a 0 mV (P > 0,05). O tramadol, (+)- e (-)-tramadol mudaram a inativação de estado estacionário de I Ca-L para potenciais de membrana mais negativos. Além disso, tramadol e (+)-tramadol alteraram significativamente a curva de recuperação dependente de tempo da I Ca-L para a direita e reduziram a recuperação de I Ca-L da inativação. A constante de tempo foi aumentada de 175,6 ± 18,6 a 305,0 ± 32,9 ms (P < 0,01) para o tramadol e de 248,1 ± 28,1 ms para 359,0 ± 23,8 ms (P < 0,05) para o (+)-tramadol. O agonista do receptor µ-opioide (DAMGO) não tem nenhum efeito na I Ca-L. CONCLUSÃO: A inibição da I Ca-L induzida por tramadol e seus enantiômeros não teve relação com a ativação de receptores opioides e poderia explicar, pelo menos em parte, seu efeito inotrópico negativo cardíaco.

BACKGROUND: Tramadol is a centrally acting analgesic, whose mechanism of action involves opioid-receptor activation. Previously, we have shown that tramadol and its enantiomers had a negative inotropic effect on the papillary muscle in which the (+)-enantiomer is more potent than (-)- and (±)-tramadol. OBJECTIVE: In this study, we investigated the effects of tramadol and its enantiomers on L-type calcium current (I Ca-L). RESULTS: Tramadol (200 µM) reduced the peak amplitude of I Ca-L at potentials from 0 to +50 mV. At 0 mV, I Ca-L was reduced by 33.7 ± 7.2 percent. (+)- and (-)-tramadol (200 µM) produced a similar inhibition of I Ca-L, in which the peak amplitude was reduced by 64.4 ± 2.8 percent and 68.9 ± 5.8 percent, respectively at 0 mV (p > 0.05). Tramadol, (+)- and (-)-tramadol shifted the steady-state inactivation of I Ca-L to more negative membrane potentials. Also, tramadol and (+)-tramadol markedly shifted the time-dependent recovery curve of I Ca-L to the right and slowed down the recovery of I Ca-L from inactivation. The time constant was increased from 175.6 ± 18.6 to 305.0 ± 32.9 ms (p < 0.01) for tramadol and from 248.1 ± 28.1 ms to 359.0 ± 23.8 ms (p < 0.05) for (+)-tramadol. The agonist of µ-opioid receptor DAMGO had no effect on the I Ca-L. CONCLUSION: The inhibition of I Ca-L induced by tramadol and its enantiomers was unrelated to the activation of opioid receptors and could explain, at least in part, their negative cardiac inotropic effect.

The influence of age on bupivacaine cardiotoxicity.

BACKGROUND: The susceptibility of children and newborns to cardiotoxicity from racemic bupivacaine, RS(±)-bupivacaine, is controversial. Some studies indicate that newborns can sustain higher bupivacaine plasma levels than adults, without severe toxicity. In this study, we compared the influence of age on cardiotoxicity from RS(±)-bupivacaine and S(-)-bupivacaine in rats. The effects of these local anesthetics (LAs) on the regulation of intracellular Ca(2+) concentrations in cardiac fibers were also investigated. METHODS: The lethal dose was determined in ventilated male Wistar rats at 2, 4, 8, and 16 weeks of age by monitoring when cardiac electrical activity stopped after infusion of RS(±)-bupivacaine and S(-)-bupivacaine (4 mg · kg(-1) · min(-1)). The effects on cardiac muscle contraction were investigated by in vitro measurement of papillary muscle twitches in the presence and absence of RS(±)-bupivacaine or S(-)-bupivacaine. Skinned ventricular fibers were used to investigate the intracellular effects on Ca(2+) regulation induced by both LAs. RESULTS: The lethal dose for RS(±)-bupivacaine and S(-)-bupivacaine in 2-week-old animals (46.0 ± 5.2 and 91.3 ± 4.9 mg · kg(-1), respectively) was higher than in 16-week-old animals (22.7 ± 1.3 and 22.0 ± 2.7 mg · kg(-1), respectively). Papillary muscle twitches were reduced in a dose-dependent manner, with significant difference between young and adult hearts. In adults, the muscle twitches were reduced to 8.6% ± 0.8% of control by RS(±)-bupivacaine, and to 18.1% ± 2.7% of control by S(-)-bupivacaine (100 µM). S(-)-bupivacaine had a positive inotropic effect at <10 µM, but only in 2-week-old animals. In chemically skinned ventricular fibers, RS(±)-bupivacaine and S(-)-bupivacaine induced similar increases in Ca(2+) release from the sarcoplasmic reticulum (SR) preactivated with caffeine (1 mM), and this effect was greater in younger rats than adults. In 16-week-old rats, caffeine-induced tension was 53.9% ± 1.7% of the maximal fiber response with RS(±)-bupivacaine, and 54.1% ± 3.2% with S(-)-bupivacaine. The caffeine response in 2-week-old rats was 81.1% ± 3.7% of the maximal response with RS(±)-bupivacaine, and 78.1% ± 4.5% with S(-)-bupivacaine. The Ca(2+) sensitivity of contractile proteins was equally increased at both ages tested, with RS(±)-bupivacaine or S(-)-bupivacaine. Ca(2+) uptake from the SR was not altered by the LA or by age. CONCLUSIONS: Differences in the mechanisms for regulating intracellular SR Ca(2+) may contribute to the decreased susceptibility of young animals to cardiodepression induced by RS(±)-bupivacaine and S(-)-bupivacaine.

The synergistic interaction between morphine and maprotiline after intrathecal injection in rats.

BACKGROUND: Antidepressant drugs act as potent inhibitors of norepinephrine and/or serotonin reuptake and are widely used with opioids for the treatment of chronic pain. The mechanism of this increased analgesic action is unclear. We compared the antinociceptive effects of the intrathecal administration of morphine with that of a nonselective (amitriptyline) or selective (maprotiline or citalopram) antidepressant drug using the thermal withdrawal test in rats. We also investigated the possible mechanisms involved in the interactions of these drugs. METHODS: Male Wistar rats were anesthetized with sevoflurane and administered morphine and antidepressant drugs, or saline, through intrathecal injection. The antinociceptive effect was evaluated using the thermal withdrawal test before and after drug administration. The time for the withdrawal reaction was expressed as percentage of maximum possible effect (MPE). Animals were also pretreated with yohimbine (a nonselective alpha2-adrenergic antagonist) and naloxone (a nonselective opioid antagonist) for mechanism of action studies. Pharmacologic interaction was evaluated using isobolographic analysis of simultaneous administration of fixed proportions of maprotiline and morphine. RESULTS: Single intrathecal administration of morphine (2 microg), amitriptiline (125 microg), citalopram (144 microg), and maprotiline (1.25 microg) produced 51.6% +/- 8.9%, 10.3% +/- 3.2%, 33.8% +/- 5.2%, and 48.5% +/- 9.2% MPE, respectively. The antinociceptive effect of morphine was increased when combined with amitriptyline (91.3% +/- 4.6% MPE) and maprotiline (86.9% +/- 9.2% MPE) but not with citalopram (40.6% +/- 4.6% MPE). Coadministration of maprotiline increased the antinociceptive duration of morphine by 4-fold (from 120 to 480 min), which was reversed by pretreatment with the alpha-2 adrenoceptor inhibitor, yohimbine, and the mu-type opioid receptor antagonist, naloxone. Isobolographic analysis demonstrated a synergistic interaction between morphine and maprotiline. CONCLUSIONS: Selective norepinephrine reuptake inhibitors can significantly increase the intensity and duration of morphine antinociceptive activity via both alpha(2)-adrenergic and opioid receptors. This interaction was not observed with the selective serotonin inhibitor, citalopram.

Comparative effects of tramadol on vascular reactivity in normotensive and spontaneously hypertensive rats.

The aim of the present study was to determine the effects of tramadol on vascular reactivity in aortic rings from Wistar-Kyoto (WKY) rats and spontaneously hypertensive rats (SHR). Aortic rings, with or without endothelium, were obtained from male WKY rats and SHR (15-20 weeks old) and prepared for isometric tension recording. Aortic rings were precontracted with phenylephrine (10 micromol/L) or 40 mmol/L KCl and then exposed to cumulative concentrations of tramadol (0.1-1 mmol/L). Tramadol produced a concentration-dependent relaxation of precontracted aortic rings from WKY rats and SHR, which was not dependent on functional endothelium. Vascular relaxation was significantly greater in rings from SHR than WKY rats. The concentration of tramadol necessary to produce a 50% reduction of the maximal contraction to phenylephrine (IC(50)) in rings with and without endothelium from SHR was 0.47 +/- 0.08 and 0.44 +/- 0.03 mmol/L, respectively (P = 0.76). Tramadol attenuated the contracture elicited by Ca2+ in depolarized tissue, suggesting that it may inhibit L-type Ca2+ channels. However, pretreatment with nicardipine (1 micromol/L) prevented the relaxation induced by tramadol in aortic rings from WKY rats and partially reduced its inhibitory effect in aortic rings from SHR. 6. Pretreatment of endothelium-denuded aorta with glybenclamide (3 micromol/L), 4-aminopyridine (3 mmol/L), tetraethylammonium (3 mmol/L) and naloxone (100 micromol/L) did not affect tramadol-induced vasodilation of aortic rings from either WKY rats or SHR. Intravenous administration of tramadol (10 mg/kg) to conscious SHR significantly reduced both systolic and diastolic blood pressure from 171.4 +/- 5.3 to 129.3 +/- 5.3 (P = 0.002) and from 125.0 +/- 6.5 to 57.8 +/- 8.9 mmHg (P = 0.003), respectively.

Vasodilatory activity of novel carbamate derivatives of isatin.

Isatin (1H-indole-2,3 dione) is an endogenous compound that may act as a physiological regulator of muscle contraction by reducing cGMP production by inhibition of guanylyl cyclase (GC) activity. Intracellular cGMP levels can regulate the contractile response of smooth muscle. Therefore, in the present study we investigated the effects of seven novel carbamate derivatives of isatin, namely C1-C7, on the contractility of aortic rings from Wistar rats. Carbamates C1 and C6 most effectively promoted endothelium-dependent relaxation of aortic rings pretreated with 10 micromol/L phenylephrine (PE) to induce contraction. The concentration of the C1 and C6 carbamates necessary to reduce PE-induced aortic contraction by 50% (IC(50)) was 5.6 +/- 1.0 and 48.4 +/- 3.4 micromol/L, respectively. Carbamate derivative-induced vasodilation required an intact endothelium, which is responsible for nitric oxide (NO) release. Pretreatment of rings with 100 micromol/L naloxone or 10 micromol/L atropine prevented the C1- and C6-mediated vascular relaxation, indicating that the vasodilatory activity was dependent on the activation of opioid or muscarinic receptors, respectively. The results of our studies provide insights into the role of novel carbamates in the regulation of vascular tone. Carbamates could stimulate NO synthesis, which induces vasodilation primarily by stimulation of GC and cGMP production. Taken together, our findings suggest that carbamate derivative-induced vasodilation may be considered an alternative treatment for primary and/or secondary hypertension.

Sedative-hypnotic profile of novel isatin ketals.

Isatin (1H-indol-2,3-dione) is an endogenous compound found in many tissues and fluids. Isatin and its derivatives exert pharmacological effects on the central nervous system, including anxiogenic, sedative and anticonvulsant activities. Two new groups of isatin derivatives were synthesized (nine dioxolane ketals and nine dioxane ketals) and studied for their sedative, hypnotic and anesthetic effects using pentobarbital-induced sleeping time, locomotor activity evaluation and intravenous infusion. The dioxolane ketals were more potent than dioxane ketals for inducing sedative-hypnotic states, causing up to a three-fold increase in pentobarbital hypnosis. The dioxolane ketals produced sedation, demonstrated by decreased spontaneous locomotor activity in an open field. Hypnosis and anesthesia were observed during intravenous infusion of 5'-chlorospiro-[1,3-dioxolane-2,3'-indolin]-2'-one (T3) in conscious Wistar rats. Complete recovery from hypnosis and anesthesia required 39.1+/-7.3 and 6.8+/-2.4 min, respectively. Changes in hemodynamic parameters after infusion of 5.0 mg/kg/min were minimal. These findings suggest that these new isatin derivatives represent potential candidates for the development of new drugs that act on the central nervous system and may lead to a new centrally acting anesthetic with no toxic effects on the cardiovascular or respiratory systems.

Regulation of intracellular calcium by bupivacaine isomers in cardiac myocytes from Wistar rats.

In this study we investigated the effects of a racemic mixture of bupivacaine (RS(+/-)bupivacaine) and its isomers (S(-)bupivacaine and R(+)bupivacaine) on the Ca2+ handling by ventricular myocytes from Wistar rats. Single ventricular myocytes were enzymatically isolated and loaded with the fluorescent Ca2+ indicator fura 2-am to estimate intracellular Ca2+ concentration during contraction and relaxation cycles. S(-)bupivacaine (10 muM) significantly increased peak amplitude and the rate of increase of Ca2+ transients in 155% +/- 54% (P < 0.05) and 194% +/- 94% (P < 0.01) of control. However, exposure to R(+)bupivacaine had no effect on either peak amplitude or rate of increase at any concentration tested. Saponin-skinned ventricular fibers were used to investigate the effect of bupivacaine on the intracellular Ca2+ regulation by sarcoplasmic reticulum (SR) and on the Ca2+ sensitivity of contractile system. S(-), R(+), and RS(+/-)bupivacaine induced Ca2+ release from SR (P < 0.01). In SR-disrupted skinned ventricular cells, bupivacaine and its isomers (5 mM) increased the sensitivity of contractile system to Ca(2+). S(-), RS(+/-), and R(+)bupivacaine significantly increased pCa50 from 5.8 +/- 0.1, 5.8 +/- 0.1, and 5.8 +/- 0.1, to 6.1 +/- 0.1 (P < 0.05), 6.0 +/- 0.1 (P < 0.05), and 6.1 +/- 0.1 (P < 0.05). Ca2+ release from SR through RyR2 activation could explain the increase of Ca2+ transients in cardiac cells. Increased intracellular Ca2+ in cardiac myocytes display a stereoselectivity to S(-)bupivacaine.