RESUMO
Ca2+ channels with a CaV1.3 pore-forming α1 subunit have been implicated in both neurodegenerative and neuropsychiatric disorders, motivating the development of selective and potent inhibitors of CaV1.3 versus CaV1.2 channels, the calcium channels implicated in hypertensive disorders. We have previously identified pyrimidine-2,4,6-triones (PYTs) that preferentially inhibit CaV1.3 channels, but the structural determinants of their interaction with the channel have not been identified, impeding their development into drugs. By a combination of biochemical, computational, and molecular biological approaches, it was found that PYTs bind to the dihydropyridine (DHP) binding pocket of the CaV1.3 subunit, establishing them as negative allosteric modulators of channel gating. Site-directed mutagenesis, based on homology models of CaV1.3 and CaV1.2 channels, revealed that a single amino acid residue within the DHP binding pocket (M1078) is responsible for the selectivity of PYTs for CaV1.3 over CaV1.2. In addition to providing direction for chemical optimization, these results suggest that, like dihydropyridines, PYTs have pharmacological features that could make them of broad clinical utility.
Assuntos
Bloqueadores dos Canais de Cálcio/metabolismo , Canais de Cálcio Tipo L/metabolismo , Pirimidinonas/metabolismo , Regulação Alostérica , Sítio Alostérico , Animais , Cálcio/metabolismo , Canais de Cálcio Tipo L/química , Canais de Cálcio Tipo L/genética , Neurônios Dopaminérgicos/efeitos dos fármacos , Células HEK293 , Humanos , Camundongos , Simulação de Acoplamento Molecular , Mutagênese Sítio-Dirigida , Mutação , Ligação Proteica , Coelhos , RatosRESUMO
In the last decade, scientific and clinical interest in the pedunculopontine nucleus (PPN) has grown dramatically. This growth is largely a consequence of experimental work demonstrating its connection to the control of gait and of clinical work implicating PPN pathology in levodopa-insensitive gait symptoms of Parkinson's disease (PD). In addition, the development of optogenetic and chemogenetic approaches has made experimental analysis of PPN circuitry and function more tractable. In this brief review, recent findings in the field linking PPN to the basal ganglia and PD are summarized; in addition, an attempt is made to identify key gaps in our understanding and challenges this field faces in moving forward.
Assuntos
Neurônios Colinérgicos/patologia , Marcha , Doença de Parkinson/patologia , Núcleo Tegmental Pedunculopontino/patologia , HumanosRESUMO
Burst spiking in substantia nigra pars compacta (SNc) dopaminergic neurons is a key signaling event in the circuitry controlling goal-directed behavior. It is widely believed that this spiking mode depends upon an interaction between synaptic activation of N-methyl-D-aspartate receptors (NMDARs) and intrinsic oscillatory mechanisms. However, the role of specific neural networks in burst generation has not been defined. To begin filling this gap, SNc glutamatergic synapses arising from pedunculopotine nucleus (PPN) neurons were characterized using optical and electrophysiological approaches. These synapses were localized exclusively on the soma and proximal dendrites, placing them in a good location to influence spike generation. Indeed, optogenetic stimulation of PPN axons reliably evoked spiking in SNc dopaminergic neurons. Moreover, burst stimulation of PPN axons was faithfully followed, even in the presence of NMDAR antagonists. Thus, PPN-evoked burst spiking of SNc dopaminergic neurons in vivo may not only be extrinsically triggered, but extrinsically patterned as well.
Assuntos
Potenciais de Ação , Neurônios Dopaminérgicos/fisiologia , Rede Nervosa/fisiologia , Núcleo Tegmental Pedunculopontino/fisiologia , Substância Negra/fisiologia , Animais , Camundongos Endogâmicos C57BL , OptogenéticaRESUMO
Dopaminergic (DA) neurons in the substantia nigra pars compacta (SNc) are richly innervated by GABAergic neurons. The postsynaptic effects of GABA on SNc DA neurons are mediated by a mixture of GABAA and GABAB receptors. Although activation of GABAA receptors inhibits spike generation, the consequences of GABAB receptor activation are less well characterized. To help fill this gap, perforated patch recordings were made from young adult mouse SNc DA neurons. Sustained stimulation of GABAB receptors hyperpolarized SNc DA neurons, as previously described. However, transient stimulation of GABAB receptors by optical uncaging of GABA did not; rather, it reduced the opening of small-conductance, calcium-activated K+ (SK) channels and increased the irregularity of spiking. This modulation was attributable to inhibition of adenylyl cyclase and protein kinase A. Thus, because suppression of SK channel activity increases the probability of burst spiking, transient co-activation of GABAA and GABAB receptors could promote a pause-burst pattern of spiking.
Assuntos
Neurônios Dopaminérgicos/metabolismo , Ativação do Canal Iônico/efeitos dos fármacos , Parte Compacta da Substância Negra/metabolismo , Receptores de GABA-B/metabolismo , Canais de Potássio Ativados por Cálcio de Condutância Baixa/metabolismo , Ácido gama-Aminobutírico/farmacologia , Adenilil Ciclases/metabolismo , Animais , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Feminino , Neurônios GABAérgicos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Técnicas de Patch-Clamp , Receptores de GABA-A/metabolismoRESUMO
Loss of noradrenergic locus coeruleus (LC) neurons is a prominent feature of aging-related neurodegenerative diseases, such as Parkinson's disease (PD). The basis of this vulnerability is not understood. To explore possible physiological determinants, we studied LC neurons using electrophysiological and optical approaches in ex vivo mouse brain slices. We found that autonomous activity in LC neurons was accompanied by oscillations in dendritic Ca(2+) concentration that were attributable to the opening of L-type Ca(2+) channels. This oscillation elevated mitochondrial oxidant stress and was attenuated by inhibition of nitric oxide synthase. The relationship between activity and stress was malleable, as arousal and carbon dioxide increased the spike rate but differentially affected mitochondrial oxidant stress. Oxidant stress was also increased in an animal model of PD. Thus, our results point to activity-dependent Ca(2+) entry and a resulting mitochondrial oxidant stress as factors contributing to the vulnerability of LC neurons.
Assuntos
Dendritos/enzimologia , Locus Cerúleo/enzimologia , Mitocôndrias/enzimologia , Óxido Nítrico Sintase/fisiologia , Estresse Oxidativo/fisiologia , Animais , Canais de Cálcio Tipo L/fisiologia , Ativação Enzimática/fisiologia , Locus Cerúleo/citologia , Locus Cerúleo/metabolismo , Masculino , Potencial da Membrana Mitocondrial/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mitocôndrias/metabolismoRESUMO
Anticholinesterases are the most common treatment for Alzheimer's disease, and, in recent years, a new group of cholinesterase inhibitors (i.e. rivastigmine, galantamine, and donepezil) has become available. Although these drugs improve cognitive symptoms, they also can induce or exacerbate parkinsonian symptoms, including tremor. The present studies were conducted to determine if galantamine induces tremulous jaw movements, a rodent model of parkinsonian tremor, and to investigate whether these oral motor impairments can be reversed by co-administration of adenosine A(2A) antagonists. The first experiment demonstrated that systemic injections of galantamine (0.75-6.0 mg/kg I.P.) induced a dose-related increase in tremulous jaw movements in rats. In a second study, co-administration of the muscarinic antagonist scopolamine (0.0156-0.25 mg/kg I.P.) produced a dose dependent suppression of tremulous jaw movements induced by a 3.0 mg/kg dose of galantamine, indicating that galantamine induces these tremulous oral movements through actions on muscarinic acetylcholine receptors. In two additional studies, analyses of freeze-frame video and electromyographic activity recorded from the lateral temporalis muscle indicated that the local frequency of these galantamine-induced jaw movements occurs in the 3-7 Hz frequency range that is characteristic of parkinsonian tremor. In the final experiment, the adenosine A(2A) antagonist MSX-3 significantly attenuated the tremulous jaw movements induced by the 3.0mg/kg dose of galantamine, which is consistent with the hypothesis that co-administration of adenosine A(2A) antagonists may be beneficial in reducing parkinsonian motor impairments induced by anticholinesterase treatment.
Assuntos
Doença de Alzheimer/tratamento farmacológico , Colinérgicos/toxicidade , Modelos Animais de Doenças , Galantamina/toxicidade , Transtornos Parkinsonianos/induzido quimicamente , Tremor/induzido quimicamente , Doença de Alzheimer/fisiopatologia , Animais , Relação Dose-Resposta a Droga , Eletromiografia/métodos , Arcada Osseodentária/efeitos dos fármacos , Arcada Osseodentária/fisiopatologia , Masculino , Transtornos Parkinsonianos/fisiopatologia , Ratos , Ratos Sprague-Dawley , Resultado do Tratamento , Tremor/fisiopatologiaRESUMO
Forebrain dopamine (DA) is a critical component of the brain circuitry regulating behavioral activation. Adenosine A(2A) antagonists reverse many of the behavioral effects of DA antagonists, and A(2A) receptors are co-localized with D(2) receptors on striatal medium spiny neurons. The present work was undertaken to determine if the ability of an A(2A) antagonist, a non-selective adenosine antagonist, or an A(1) antagonist to reverse the locomotor effects of DA blockade in rats differed depending upon whether D(1) or D(2) family receptors were being antagonized. The adenosine antagonists MSX-3, caffeine, DPCPX and CPT were studied for their ability to reverse the locomotor suppression induced by the D(1) antagonist SCH 39166 (ecopipam) and the D(2) antagonist eticlopride. The D(1) and D(2) antagonists suppressed locomotion in all experiments. The adenosine A(2A) receptor antagonist MSX-3 (0.5-2.0 mg/kg IP) significantly reversed the suppression of locomotion induced by eticlopride. The non-selective adenosine antagonist caffeine (5.0-20.0 mg/kg IP) also reversed the effect of eticlopride, though the effect was not as robust as that seen with MSX-3. The adenosine A(1) antagonists DPCPX (0.375-1.5 mg/kg) and CPT (3.0-12.0 mg/kg IP) were unable to reverse the locomotor impairment elicited by eticlopride. Furthermore, the attenuation of locomotion induced by the D(1) antagonist could only be reversed by the highest dose of MSX-3, but not by caffeine, DPCPX or CPT. DA and adenosine receptor antagonists interact in the regulation of locomotor activation, but the nature of this interaction appears to depend upon the receptor selectivity profiles of the specific drugs being tested.
Assuntos
Antagonistas de Dopamina/farmacologia , Atividade Motora/efeitos dos fármacos , Prosencéfalo/efeitos dos fármacos , Antagonistas de Receptores Purinérgicos P1 , Receptores Dopaminérgicos/efeitos dos fármacos , Análise de Variância , Animais , Comportamento Animal/efeitos dos fármacos , Relação Dose-Resposta a Droga , Interações Medicamentosas , Masculino , Ratos , Ratos Sprague-Dawley , Receptores Dopaminérgicos/metabolismo , Receptores Purinérgicos P1/classificação , Receptores Purinérgicos P1/metabolismoRESUMO
Tremulous jaw movements in rats, which can be induced by dopamine (DA) antagonists, DA depletion, and cholinomimetics, have served as a useful model for studies of tremor. Although adenosine A(2A) antagonists can reduce the tremulous jaw movements induced by DA antagonists and DA depletion, there are conflicting reports about the interaction between adenosine antagonists and cholinomimetic drugs. The present studies investigated the ability of adenosine antagonists to reverse the tremorogenic effect of the muscarinic agonist pilocarpine. While the adenosine A(2A) antagonist MSX-3 was incapable of reversing the tremulous jaw movements induced by the 4.0mg/kg dose of pilocarpine, both MSX-3 and the adenosine A(2A) antagonist SCH58261 reversed the tremulous jaw movements elicited by 0.5mg/kg pilocarpine. Systemic administration of the adenosine A(1) antagonist DPCPX failed to reverse the tremulous jaw movements induced by either an acute 0.5mg/kg dose of the cholinomimetic pilocarpine or the DA D2 antagonist pimozide, indicating that the tremorolytic effects of adenosine antagonists may be receptor subtype specific. Behaviorally active doses of MSX-3 and SCH 58261 showed substantial in vivo occupancy of A(2A) receptors, but DPCPX did not. The results of these studies support the use of adenosine A(2A) antagonists for the treatment of tremor.