RESUMEN
Mephedrone (4-methylmethcathinone) has been found in several over-the-counter products that are abused by humans, but very little is known about its behavioral effects and abuse liability. The present study examined the effects of mephedrone (1-10 mg/kg) on learning in female rats, as well as its interaction with the ovarian hormone estradiol. More specifically, female rats were trained to respond under a multiple schedule of repeated acquisition and performance of response sequences and then ovariectomized. Following ovariectomy, mephedrone dose-effect curves were obtained during periods of 17ß-estradiol administration and periods without estradiol administration. Unlike mephedrone, which was administered acutely (i.p.) before the experimental sessions, 17ß-estradiol was administered via subcutaneous Silastic capsules containing 25% 17ß-estradiol and 75% cholesterol. In general, mephedrone produced dose-dependent rate-decreasing and error-increasing effects in the acquisition and performance components of the schedule in all subjects. However, when estradiol was present, three of the four rats were more sensitive to the rate-decreasing effects of mephedrone, and all of the subjects were more sensitive to its error-increasing effects. These data indicate that estradiol can potentiate the disruptive effects of mephedrone on both the acquisition and performance of complex behavior in female rats.
Asunto(s)
Condicionamiento Operante/efectos de los fármacos , Estradiol/farmacología , Drogas Ilícitas/farmacología , Metanfetamina/análogos & derivados , Animales , Relación Dosis-Respuesta a Droga , Sinergismo Farmacológico , Femenino , Metanfetamina/farmacología , Ovariectomía , Ratas , Ratas Long-EvansRESUMEN
RATIONALE: Exceedingly little experimental research exists on the popular recreational drug mephedrone (4-methylmethcathinone) despite clinical reports concerning its behavioral and cardiovascular toxicity. OBJECTIVES: To characterize mephedrone preclinically by examining its capacity to (1) serve as a discriminative stimulus, (2) disrupt the acquisition of response sequences, and (3) disrupt mean arterial pressure (MAP) and heart rate (HR). METHODS AND RESULTS: In one group of subjects that reliably discriminated 3.2 mg/kg of mephedrone from saline (n = 9), substitution tests indicated that stimulants (cocaine, MDMA, and methamphetamine) more closely approximated the mephedrone discriminative stimulus than non-stimulants (fenfluramine, morphine, and phencyclidine), although none fully substituted. In a second group (n = 6), mephedrone (0.56-10 mg/kg, i.p.) dose-dependently decreased response rate and increased errors in both components of a procedure in which subjects either acquired a new response sequence each session (repeated acquisition) or completed the same response sequence each session (performance). Finally, in a third group (n = 12), radio telemetry probes were used to measure the changes in MAP and HR elicited by mephedrone and then compared them to a known stimulant, methamphetamine. In these studies, mephedrone (0.01-9 mg/kg, i.v.) elicited increases in MAP and HR that were very similar to those elicited by methamphetamine (0.01-9 mg/kg, i.v.). The tachycardia and pressor responses to mephedrone (3 mg/kg) were blocked by the ß-blocker atenolol (1 mg/kg, i.v.) and the α1, α2-blocker phentolamine (3 mg/kg, i.v.), respectively. CONCLUSIONS: Mephedrone produces behavioral and cardiovascular responses that are similar to other stimulants; however, differences from the classical stimulants were also apparent.
Asunto(s)
Conducta Animal/efectos de los fármacos , Presión Sanguínea/efectos de los fármacos , Frecuencia Cardíaca/efectos de los fármacos , Drogas Ilícitas/toxicidad , Metanfetamina/análogos & derivados , Análisis de Varianza , Animales , Condicionamiento Operante/efectos de los fármacos , Aprendizaje Discriminativo/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Masculino , Metanfetamina/toxicidad , Ratas , Ratas Long-Evans , Ratas Sprague-DawleyRESUMEN
The relationship between cerebellar dysfunction, motor symptoms, and neuronal loss in the inherited ataxias, including the polyglutamine disease spinocerebellar ataxia type 3 (SCA3), remains poorly understood. We demonstrate that before neurodegeneration, Purkinje neurons in a mouse model of SCA3 exhibit increased intrinsic excitability resulting in depolarization block and the loss of the ability to sustain spontaneous repetitive firing. These alterations in intrinsic firing are associated with increased inactivation of voltage-activated potassium currents. Administration of an activator of calcium-activated potassium channels, SKA-31, partially corrects abnormal Purkinje cell firing and improves motor function in SCA3 mice. Finally, expression of the disease protein, ataxin-3, in transfected cells increases the inactivation of Kv3.1 channels and shifts the activation of Kv1.2 channels to more depolarized potentials. Our results suggest that in SCA3, early Purkinje neuron dysfunction is associated with altered physiology of voltage-activated potassium channels. We further suggest that the observed changes in Purkinje neuron physiology contribute to disease pathogenesis, underlie at least some motor symptoms, and represent a promising therapeutic target in SCA3.
Asunto(s)
Cerebelo/fisiopatología , Enfermedad de Machado-Joseph/fisiopatología , Péptidos/fisiología , Animales , Benzotiazoles , Western Blotting , Muerte Celular/fisiología , Línea Celular , Venenos Elapídicos/farmacología , Fenómenos Electrofisiológicos , Humanos , Inmunohistoquímica , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Ratones Transgénicos , Trastornos del Movimiento/fisiopatología , Degeneración Nerviosa/patología , Conducción Nerviosa/fisiología , Neuronas/fisiología , Técnicas de Placa-Clamp , Canales de Potasio Calcio-Activados/fisiología , Células de Purkinje/fisiología , Canales de Potasio de la Superfamilia Shaker/metabolismoRESUMEN
Small-conductance (KCa2.1-2.3) and intermediate-conductance (KCa3.1) calcium-activated K(+) channels are critically involved in modulating calcium-signaling cascades and membrane potential in both excitable and nonexcitable cells. Activators of these channels constitute useful pharmacological tools and potential new drugs for the treatment of ataxia, epilepsy, and hypertension. Here, we used the neuroprotectant riluzole as a template for the design of KCa2/3 channel activators that are potent enough for in vivo studies. Of a library of 41 benzothiazoles, we identified 2 compounds, anthra[2,1-d]thiazol-2-ylamine (SKA-20) and naphtho[1,2-d]thiazol-2-ylamine (SKA-31), which are 10 to 20 times more potent than riluzole and activate KCa2.1 with EC(50) values of 430 nM and 2.9 microM, KCa2.2 with an EC(50) value of 1.9 microM, KCa2.3 with EC(50) values of 1.2 and 2.9 microM, and KCa3.1 with EC(50) values of 115 and 260 nM. Likewise, SKA-20 and SKA-31 activated native KCa2.3 and KCa3.1 channels in murine endothelial cells, and the more "drug-like" SKA-31 (half-life of 12 h) potentiated endothelium-derived hyperpolarizing factor-mediated dilations of carotid arteries from KCa3.1(+/+) mice but not from KCa3.1(-/-) mice. Administration of 10 and 30 mg/kg SKA-31 lowered mean arterial blood pressure by 4 and 6 mm Hg in normotensive mice and by 12 mm Hg in angiotensin-II-induced hypertension. These effects were absent in KCa3.1-deficient mice. In conclusion, with SKA-31, we have designed a new pharmacological tool to define the functional role of the KCa2/3 channel activation in vivo. The blood pressure-lowering effect of SKA-31 suggests KCa3.1 channel activation as a new therapeutic principle for the treatment of hypertension.
Asunto(s)
Benzotiazoles/farmacología , Factores Biológicos/metabolismo , Presión Sanguínea/efectos de los fármacos , Endotelio/efectos de los fármacos , Canales de Potasio de Conductancia Intermedia Activados por el Calcio/metabolismo , Animales , Ratones , Canales de Potasio/metabolismoRESUMEN
Calcium-activated potassium channels modulate calcium signaling cascades and membrane potential in both excitable and non-excitable cells. In this article we will review the physiological properties, the structure activity relationships of the existing peptide and small molecule modulators and the therapeutic importance of the three small-conductance channels KCa2.1-KCa2.3 (a.k.a. SK1-SK3) and the intermediate-conductance channel KCa3.1 (a.k.a. IKCa1). The apamin-sensitive KCa2 channels contribute to the medium afterhyperpolarization and are crucial regulators of neuronal excitability. Based on behavioral studies with apamin and on observations made in several transgenic mouse models, KCa2 channels have been proposed as targets for the treatment of ataxia, epilepsy, memory disorders and possibly schizophrenia and Parkinson's disease. In contrast, KCa3.1 channels are found in lymphocytes, erythrocytes, fibroblasts, proliferating vascular smooth muscle cells, vascular endothelium and intestinal and airway epithelia and are therefore regarded as targets for various diseases involving these tissues. Since two classes of potent and selective small molecule KCa3.1 blocker, triarylmethanes and cyclohexadienes, have been identified, several of these postulates have already been validated in animal models. The triarylmethane ICA-17043 is currently in phase III clinical trials for sickle cell anemia while another triarylmethane, TRAM-34, has been shown to prevent vascular restenosis in rats and experimental autoimmune encephalomyelitis in mice. Experiments showing that a cyclohexadiene KCa3.1 blocker reduces infarct volume in a rat subdural hematoma model further suggest KCa3.1 as a target for the treatment of traumatic and possibly ischemic brain injury. Taken together KCa2 and KCa3.1 channels constitute attractive new targets for several diseases that currently have no effective therapies.
Asunto(s)
Enfermedades del Sistema Nervioso/fisiopatología , Neuronas/fisiología , Canales de Potasio Calcio-Activados/fisiología , Canales de Potasio de Pequeña Conductancia Activados por el Calcio/fisiología , Secuencia de Aminoácidos , Animales , Apamina/farmacología , Caribdotoxina/farmacología , Modelos Animales de Enfermedad , Humanos , Canales de Potasio de Conductancia Intermedia Activados por el Calcio/efectos de los fármacos , Canales de Potasio de Conductancia Intermedia Activados por el Calcio/fisiología , Masculino , Datos de Secuencia MolecularRESUMEN
The voltage-gated potassium channel Kv1.3 has been recently identified as a molecular target that allows for selective pharmacological suppression of effector memory T (T(EM)) cells without affecting the function of naïve and central memory T cells. We here investigated whether PAP-1, a small molecule Kv1.3 blocker (EC50=2 nM), could suppress allergic contact dermatitis (ACD). In a rat model of ACD, we first confirmed that the infiltrating cells in the elicitation phase are indeed CD8+ CD45RC- memory T cells with high Kv1.3 expression. In accordance with its selective effect on T(EM) cells, PAP-1 did not impair sensitization, but potently suppressed oxazolone-induced inflammation by inhibiting the infiltration of CD8+ T cells and reducing the production of the inflammatory cytokines IFN-gamma, IL-2, and IL-17 when administered intraperitoneally or orally during the elicitation phase. PAP-1 was equally effective when applied topically, demonstrating that it effectively penetrates skin. We further show that PAP-1 is not a sensitizer or an irritant and exhibits no toxicity in a 28-day toxicity study. Based on these results we propose that PAP-1 could potentially be developed into a drug for the topical treatment of inflammatory skin diseases such as psoriasis.
Asunto(s)
Linfocitos T CD8-positivos/efectos de los fármacos , Dermatitis Alérgica por Contacto/tratamiento farmacológico , Furocumarinas/farmacocinética , Memoria Inmunológica/efectos de los fármacos , Canal de Potasio Kv1.3/metabolismo , Bloqueadores de los Canales de Potasio/farmacocinética , Adyuvantes Inmunológicos , Administración Oral , Administración Tópica , Animales , Linfocitos T CD8-positivos/inmunología , Linfocitos T CD8-positivos/metabolismo , Dermatitis Alérgica por Contacto/inmunología , Oído , Edema/inducido químicamente , Edema/tratamiento farmacológico , Femenino , Furocumarinas/sangre , Furocumarinas/toxicidad , Inmunosupresores/farmacología , Inyecciones Intraperitoneales , Inyecciones Intravenosas , Interferón gamma/metabolismo , Interleucina-17/metabolismo , Antígenos Comunes de Leucocito/metabolismo , Ratones , Ratones Endogámicos BALB C , Oxazolona , Proteínas Asociadas a Pancreatitis , Bloqueadores de los Canales de Potasio/sangre , Bloqueadores de los Canales de Potasio/toxicidad , Ratas , Ratas Endogámicas Lew , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
Autoreactive memory T lymphocytes are implicated in the pathogenesis of autoimmune diseases. Here we demonstrate that disease-associated autoreactive T cells from patients with type-1 diabetes mellitus or rheumatoid arthritis (RA) are mainly CD4+ CCR7- CD45RA- effector memory T cells (T(EM) cells) with elevated Kv1.3 potassium channel expression. In contrast, T cells with other antigen specificities from these patients, or autoreactive T cells from healthy individuals and disease controls, express low levels of Kv1.3 and are predominantly naïve or central-memory (T(CM)) cells. In T(EM) cells, Kv1.3 traffics to the immunological synapse during antigen presentation where it colocalizes with Kvbeta2, SAP97, ZIP, p56(lck), and CD4. Although Kv1.3 inhibitors [ShK(L5)-amide (SL5) and PAP1] do not prevent immunological synapse formation, they suppress Ca2+-signaling, cytokine production, and proliferation of autoantigen-specific T(EM) cells at pharmacologically relevant concentrations while sparing other classes of T cells. Kv1.3 inhibitors ameliorate pristane-induced arthritis in rats and reduce the incidence of experimental autoimmune diabetes in diabetes-prone (DP-BB/W) rats. Repeated dosing with Kv1.3 inhibitors in rats has not revealed systemic toxicity. Further development of Kv1.3 blockers for autoimmune disease therapy is warranted.
Asunto(s)
Artritis Reumatoide/inmunología , Artritis Reumatoide/metabolismo , Diabetes Mellitus Tipo 1/inmunología , Diabetes Mellitus Tipo 1/metabolismo , Canal de Potasio Kv1.3/metabolismo , Linfocitos T/inmunología , Linfocitos T/metabolismo , Animales , Artritis Reumatoide/tratamiento farmacológico , Artritis Reumatoide/patología , Diabetes Mellitus Tipo 1/tratamiento farmacológico , Diabetes Mellitus Tipo 1/patología , Modelos Animales de Enfermedad , Electrofisiología , Femenino , Humanos , Canal de Potasio Kv1.3/antagonistas & inhibidores , Proteínas Asociadas a Pancreatitis , Técnicas de Placa-Clamp , Bloqueadores de los Canales de Potasio/farmacología , Ratas , Receptores CCR7 , Receptores de Quimiocina/metabolismo , Linfocitos T/patologíaRESUMEN
The lymphocyte K+ channel Kv1.3 constitutes an attractive pharmacological target for the selective suppression of terminally differentiated effector memory T (TEM) cells in T cell-mediated autoimmune diseases, such as multiple sclerosis and type 1 diabetes. Unfortunately, none of the existing small-molecule Kv1.3 blockers is selective, and many of them, such as correolide, 4-phenyl-4-[3-(methoxyphenyl)-3-oxo-2-azapropyl]cyclohexanone, and our own compound Psora-4 inhibit the cardiac K+ channel Kv1.5. By further exploring the structure-activity relationship around Psora-4 through a combination of traditional medicinal chemistry and whole-cell patch-clamp, we identified a series of new phenoxyalkoxypsoralens that exhibit 2- to 50-fold selectivity for Kv1.3 over Kv1.5, depending on their exact substitution pattern. The most potent and "drug-like" compound of this series, 5-(4-phenoxybutoxy)psoralen (PAP-1), blocks Kv1.3 in a use-dependent manner, with a Hill coefficient of 2 and an EC50 of 2 nM, by preferentially binding to the C-type inactivated state of the channel. PAP-1 is 23-fold selective over Kv1.5, 33- to 125-fold selective over other Kv1-family channels, and 500- to 7500-fold selective over Kv2.1, Kv3.1, Kv3.2, Kv4.2, HERG, calcium-activated K+ channels, Na+,Ca2+, and Cl- channels. PAP-1 does not exhibit cytotoxic or phototoxic effects, is negative in the Ames test, and affects cytochrome P450-dependent enzymes only at micromolar concentrations. PAP-1 potently inhibits the proliferation of human TEM cells and suppresses delayed type hypersensitivity, a TEM cell-mediated reaction, in rats. PAP-1 and several of its derivatives therefore constitute excellent new tools to further explore Kv1.3 as a target for immunosuppression and could potentially be developed into orally available immunomodulators.