RESUMO
PURPOSE: Phenylketonuria (PKU) is a disorder of phenylalanine (Phe) metabolism that frequently results in epilepsy if a low Phe diet was not implemented at birth. The mechanisms by which Phe affects the brain are poorly understood. METHODS: Audiogenic seizures (AGS) were studied in female homozygous Pah(enu2) BTBR (PKU) mice. RESULTS: Adult PKU mice, 18-20 weeks of age, in contrast to wild-type and heterozygous counterparts, exhibited a full range of AGS. Younger PKU mice, 5-7 weeks of age, had higher serum Phe levels (2.22 +/- 0.20 mM) in comparison with the adult animals (1.72 +/- 0.05 mM) and were not susceptible to AGS. Among adult mice, animals susceptible to AGS had significantly lower serum Phe levels (1.62 +/- 0.06 mM) in comparison with those resistant to AGS (1.86 +/- 0.07 mM). Susceptibility to AGS tended to increase in the afternoon when serum Phe concentration decreased in comparison to evening and morning. Normalization of serum Phe level by instituting a low Phe diet generally prevented susceptibility to AGS within 12 h. Although return to a standard diet raised Phe levels to hyperphenylalaninemic within 12 h in animals treated with a low Phe diet for 2 weeks, more than 7 weeks were needed for a complete resumption of AGS. CONCLUSIONS: Transient decrease in Phe levels within hyperphenylalaninemic range may be a necessary condition for PKU-related seizures to occur. A low Phe diet prevents susceptibility to seizures, which can resume with the significant delay after termination of dietary treatment.
Assuntos
Epilepsia Reflexa/sangue , Epilepsia/sangue , Fenilalanina/sangue , Fenilcetonúrias/sangue , Estimulação Acústica , Fatores Etários , Animais , Ritmo Circadiano , Modelos Animais de Doenças , Epilepsia/diagnóstico , Epilepsia/genética , Epilepsia Reflexa/diagnóstico , Epilepsia Reflexa/genética , Feminino , Alimentos Formulados , Predisposição Genética para Doença , Camundongos , Camundongos Mutantes , Fenilalanina/administração & dosagem , Fenilalanina/metabolismo , Fenilcetonúrias/dietoterapia , Fenilcetonúrias/genéticaRESUMO
In previous studies, we determined that macrophage migration inhibitory factor (MIF), acting intracellularly via its intrinsic thiol-protein oxidoreductase (TPOR) activity, stimulates basal neuronal delayed-rectifier K(+) current (I(Kv)) and inhibits basal and angiotensin (ANG) II-induced increases in neuronal activity. These findings are the basis for our hypothesis that MIF is a negative regulator of ANG II actions in neurons. MIF has recently been recategorized as a member of the thioredoxin (Trx) superfamily of small proteins. In the present study we have examined whether Trx influences basal and ANG II-modulated I(Kv) in an effort to determine whether the Trx superfamily can exert a general regulatory influence over neuronal activity and the actions of ANG II. Intracellular application of Trx (0.8-80 nM) into rat hypothalamic/brain stem neurons in culture increased neuronal I(Kv), as measured by voltage-clamp recordings. This effect of Trx was abolished in the presence of the TPOR inhibitor PMX 464 (800 nM). Furthermore, the mutant protein recombinant human C32S/C35S-Trx, which lacks TPOR activity, failed to alter neuronal I(Kv). Trx applied at a concentration (0.08 nM) that does not alter basal I(Kv) abolished the inhibition of neuronal I(Kv) produced by ANG II (100 nM). Given our observation that ANG II increases Trx levels in neuronal cultures, it is possible that Trx (like MIF) has a negative regulatory role over basal and ANG II-stimulated neuronal activity via modulation of I(Kv). Moreover, these data suggest that TPOR may be a general mechanism for negatively regulating neuronal activity.
Assuntos
Angiotensina II/metabolismo , Tronco Encefálico/metabolismo , Canais de Potássio de Retificação Tardia/metabolismo , Hipotálamo/metabolismo , Ativação do Canal Iônico , Neurônios/metabolismo , Proteína Dissulfeto Redutase (Glutationa)/metabolismo , Tiorredoxinas/metabolismo , Angiotensina II/farmacologia , Animais , Animais Recém-Nascidos , Benzotiazóis/farmacologia , Tronco Encefálico/citologia , Tronco Encefálico/efeitos dos fármacos , Tronco Encefálico/enzimologia , Células Cultivadas , Cicloexanonas/farmacologia , Canais de Potássio de Retificação Tardia/efeitos dos fármacos , Relação Dose-Resposta a Droga , Hipotálamo/efeitos dos fármacos , Hipotálamo/enzimologia , Ativação do Canal Iônico/efeitos dos fármacos , Potenciais da Membrana , Neurônios/efeitos dos fármacos , Neurônios/enzimologia , Técnicas de Patch-Clamp , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Tiorredoxinas/antagonistas & inibidores , Tiorredoxinas/genética , Tiorredoxinas/farmacologia , Fatores de Tempo , Regulação para CimaRESUMO
OBJECTIVE: Despite the pathophysiological and therapeutic significance of the negative dromotropic effect of adenosine, its underlying ionic mechanism, and specifically the role of the adenosine-activated K(+) current (I(K,ADO)) is not experimentally defined. Therefore, we studied the contribution of I(K,ADO) to the negative dromotropic effect of adenosine. METHODS: Effects of adenosine on single atrioventricular nodal and left atrial myocytes from rabbits were studied using the whole cell configuration of the patch clamp technique. Complementary experiments were done in rabbit and guinea pig isolated hearts instrumented to measure the atrium-to-His bundle interval. RESULTS: In contrast to its effect in atrial myocytes, Ba(2+) selectively and completely blocked I(K,ADO) at membrane potentials from -70 to 0 mV in atrioventricular nodal myocytes and abolished the adenosine-induced leftward shift of the reversal membrane potential. Ba(2+) alone did not significantly prolong the A-H interval, but markedly attenuated the A-H interval prolongation caused by adenosine. In guinea pig heart, EC(50) values ( pD(2) +/- SEM) for adenosine-induced atrium-to-His bundle interval prolongation were 3.3 micromol/L (5.48 +/- 0.04) and 13.2 micromol/L (4.88 +/- 0.05, P < 0.001) in the absence and presence of Ba(2+), respectively. Despite species-dependent differences in sensitivities to adenosine (guinea pig > rabbit), the relative contribution of adenosine-activated K(+) current to the atrium-to-His bundle interval prolongation was nearly identical. In guinea pig hearts it ranged from 37.8 % (P = 0.013) to 72.5 % (P < 0.001) at 2 to 6 micromol/L adenosine, respectively. CONCLUSION: I(K,ADO) contributes significantly to the negative dromotropic effect of adenosine, but predominantly at relatively high concentrations of the nucleoside.