RESUMEN
BACKGROUND: Glutamatergic N-methyl-d-aspartate receptors (NMDARs) are well known for their sensitivity to ethanol (EtOH) inhibition. However, the specific manner in which EtOH inhibits channel activity and how such inhibition affects neurotransmission, and ultimately behavior, remains unclear. Replacement of phenylalanine 639 with alanine (F639A) in the GluN1 subunit reduces EtOH inhibition of recombinant NMDARs. Mice expressing this subunit show reduced EtOH-induced anxiolysis, blunted locomotor stimulation following low-dose EtOH administration, and faster recovery of motor function after moderate doses of EtOH, suggesting that cerebellar dysfunction may contribute to some of these behaviors. In the mature mouse cerebellum, NMDARs at the cerebellar climbing fiber (CF) to Purkinje cell (PC) synapse are inhibited by low concentrations of EtOH and the long-term depression (LTD) of parallel fiber (PF)-mediated currents induced by concurrent activation of PFs and CFs (PF-LTD) requires activation of EtOH-sensitive NMDARs. In this study, we examined cerebellar NMDA responses and NMDA-mediated synaptic plasticity in wild-type (WT) and GluN1(F639A) mice. METHODS: Patch-clamp electrophysiological recordings were performed in acute cerebellar slices from adult WT and GluN1(F639A) mice. NMDAR-mediated currents at the CF-PC synapse and NMDAR-dependent PF-LTD induction were compared for genotype-dependent differences. RESULTS: Stimulation of CFs evoked robust NMDA-mediated excitatory postsynaptic currents (EPSCs) in PCs that were similar in amplitude and kinetics between WT and GluN1(F639A) mice. NMDA-mediated CF-PC EPSCs in WT mice were significantly inhibited by EtOH (50 mM) while those in mutant mice were unaffected. Concurrent stimulation of CF and PF inputs induced synaptic depression of PF-PC EPSCs in both WT and mutant mice, and this depression was blocked by the NMDA antagonist DL-APV. The synaptic depression of PF-PC EPSCs in WT mice was also blocked by a low concentration of EtOH (10 mM) that had no effect on plasticity in GluN1(F639A) mice. CONCLUSIONS: These results demonstrate that inhibition of cerebellar NMDARs may be a key mechanism by which EtOH affects cerebellar-dependent behaviors.
Asunto(s)
Cerebelo/fisiología , Etanol/farmacología , Potenciales Postsinápticos Excitadores/fisiología , Depresión Sináptica a Largo Plazo/fisiología , Proteínas del Tejido Nervioso/antagonistas & inhibidores , Plasticidad Neuronal/fisiología , Receptores de N-Metil-D-Aspartato/antagonistas & inhibidores , Sinapsis/fisiología , Animales , Estimulación Encefálica Profunda , Potenciales Postsinápticos Excitadores/efectos de los fármacos , Ratones , Mutación , Proteínas del Tejido Nervioso/genética , Inhibición Neural/efectos de los fármacos , Receptores de N-Metil-D-Aspartato/genéticaRESUMEN
BACKGROUND: Studies indicate that exposure to ethanol (EtOH) during fetal development damages cerebellar Purkinje cells (PCs). PC proximal dendrites receive glutamatergic input from climbing fibers (CFs) originating at the inferior olive. CF input produces a characteristic response in PCs known as the complex spike (CS). During the first 2 weeks of life in rodents (equivalent to the human third trimester of pregnancy), CF-PC synapses undergo profound refinement. Here, we characterized the impact of EtOH exposure during this period on CF-evoked responses in PCs. METHODS: Using vapor chambers, neonatal rat pups and their mothers were exposed to air or EtOH for 4 h/d between postnatal day 2 (P2) and P12 (pup serum EtOH concentration, 0.16 g/dl). The function of CF-PC synapses was characterized using patch-clamp electrophysiological techniques in acute slices from the cerebellar vermis. Experiments were performed soon after EtOH withdrawal, when perisomatic CFs are still being eliminated (P15 to P17), and after weaning when CF dendritic translocation is almost complete (P21 to P34). RESULTS: Neither the baseline characteristics of the CS (Na(+) spike amplitude, area, coastline index, and afterhyperpolarization [AHP] amplitude) nor the type-1 metabotropic glutamate receptor (mGluR1)-mediated component of both the CS and AHP were significantly affected by EtOH exposure at P15 to P17 or P21 to P34. The mGluR1-dependent long-term depression (LTD) of CF-evoked excitatory postsynaptic currents was not significantly affected by EtOH exposure at P21 to P34. CONCLUSIONS: EtOH exposure during the third trimester equivalent neither affected basal characteristics of the CS nor CF-LTD at rat cerebellar PCs from juvenile rats.
Asunto(s)
Cerebelo/efectos de los fármacos , Etanol/farmacología , Células de Purkinje/efectos de los fármacos , Animales , Animales Recién Nacidos/fisiología , Benzoatos/farmacología , Cerebelo/fisiología , Etanol/efectos adversos , Antagonistas de Aminoácidos Excitadores/farmacología , Glicina/análogos & derivados , Glicina/farmacología , Masculino , Técnicas de Placa-Clamp , Células de Purkinje/fisiología , Ratas , Receptores de Glutamato Metabotrópico/antagonistas & inhibidores , Receptores de Glutamato Metabotrópico/fisiología , Transmisión Sináptica/efectos de los fármacosRESUMEN
The functional implications of transient receptor potential melastatin 3 (TRPM3) activation, the most recently described member of the melastatin subfamily of cation permeable TRP channels, have begun to be elucidated in recent years. The discovery of TRPM3 activation by the steroid pregnenolone sulfate (PregS) has shed new light on the physiological role of this channel. For example, TRPM3 activation enhances insulin secretion from ß pancreatic cells, induces contraction of vascular smooth muscle, and is also involved in the detection of noxious heat. Although TRPM3 expression has been detected in several regions of the developing and mature brain, little is known about the roles of TRPM3 in brain physiology. In this study, we demonstrate the abundant expression of TRPM3 steroid-sensitive channels in the developing cerebellar cortex. We also show that TRPM3-like channels are expressed at glutamatergic synapses in neonatal Purkinje cells. We recently showed that PregS potentiates spontaneous glutamate release onto neonatal Purkinje cells during a period of active glutamatergic synapse formation; we now show that this effect of PregS is mediated by TRPM3-like channels. Mefenamic acid, a recently discovered TRPM3 antagonist, blocked the effect of PregS on glutamate release. The PregS effect on glutamate release was mimicked by other TRPM3 agonists (nifedipine and epipregnanolone sulfate) but not by a TRMP3-inactive steroid (progesterone). Our findings identify TRPM3 channels as novel modulators of glutamatergic transmission in the developing brain.
Asunto(s)
Cerebelo/crecimiento & desarrollo , Cerebelo/metabolismo , Ácido Glutámico/fisiología , Neuronas/fisiología , Pregnenolona/farmacología , Transmisión Sináptica/fisiología , Canales Catiónicos TRPM/metabolismo , Secuencia de Aminoácidos , Animales , Animales Recién Nacidos , Cerebelo/efectos de los fármacos , Ácido Glutámico/metabolismo , Masculino , Datos de Secuencia Molecular , Neuronas/metabolismo , Transporte de Proteínas/genética , Células de Purkinje/efectos de los fármacos , Células de Purkinje/metabolismo , Células de Purkinje/fisiología , Ratas , Ratas Sprague-Dawley , Transmisión Sináptica/efectos de los fármacos , Canales Catiónicos TRPM/fisiologíaRESUMEN
Exposure to ethanol (EtOH) during fetal development can lead to long-lasting alterations, including deficits in fine motor skills and motor learning. Studies suggest that these are, in part, a consequence of cerebellar damage. Cerebellar granule neurons (CGNs) are the gateway of information into the cerebellar cortex. Functionally, CGNs are heavily regulated by phasic and tonic GABAergic inhibition from Golgi cell interneurons; however, the effect of EtOH exposure on the development of GABAergic transmission in immature CGNs has not been investigated. To model EtOH exposure during the 3rd trimester-equivalent of human pregnancy, neonatal pups were exposed intermittently to high levels of vaporized EtOH from postnatal day (P) 2 to P12. This exposure gradually increased pup serum EtOH concentrations (SECs) to â¼60 mM (â¼0.28 g/dl) during the 4 h of exposure. EtOH levels gradually decreased to baseline 8 h after the end of exposure. Surprisingly, basal tonic and phasic GABAergic currents in CGNs were not significantly affected by postnatal alcohol exposure (PAE). However, PAE increased δ subunit expression at P28 as detected by immunohistochemical and western blot analyses. Also, electrophysiological studies with an agonist that is highly selective for δ-containing GABA(A) receptors, 4,5,6,7-tetrahydroisoxazolo[4,5-c]pyridine-3-ol (THIP), showed an increase in THIP-induced tonic current. Behavioral studies of PAE rats did not reveal any deficits in motor coordination, except for a delay in the acquisition of the mid-air righting reflex that was apparent at P15 to P18. These findings demonstrate that repeated intermittent exposure to high levels of EtOH during the equivalent of the last trimester of human pregnancy has significant but relatively subtle effects on motor coordination and GABAergic transmission in CGNs in rats.
Asunto(s)
Depresores del Sistema Nervioso Central/toxicidad , Cerebelo/efectos de los fármacos , Discapacidades del Desarrollo/inducido químicamente , Etanol/toxicidad , Trastornos del Espectro Alcohólico Fetal/fisiopatología , Receptores de GABA-A/metabolismo , Animales , Depresores del Sistema Nervioso Central/sangre , Cerebelo/crecimiento & desarrollo , Cerebelo/fisiopatología , Discapacidades del Desarrollo/fisiopatología , Etanol/sangre , Femenino , Isoxazoles/farmacología , Potenciales de la Membrana/efectos de los fármacos , Potenciales de la Membrana/fisiología , Actividad Motora/efectos de los fármacos , Actividad Motora/fisiología , Neuronas/efectos de los fármacos , Neuronas/fisiología , Embarazo , Tercer Trimestre del Embarazo , Efectos Tardíos de la Exposición Prenatal/fisiopatología , Ratas , Ratas Sprague-DawleyRESUMEN
The cerebellum controls balance, posture, motor coordination, and cognition, and studies suggest that ethanol impairs these cerebellar functions. However, the mechanisms through which ethanol produces these effects are not fully understood. Here, we review evidence suggesting that ethanol acts, in part, by impairing synaptic plasticity mechanisms at cerebellar Purkinje neurons. We will primarily focus on recent experiments indicating that long-term depression at both parallel fiber- and climbing fiber-Purkinje cell synapses is inhibited by acute ethanol exposure. We will also discuss experimental evidence showing that chronic prenatal ethanol exposure converts long-term depression into long-term potentiation at parallel fiber-Purkinje cell synapses.