RESUMEN
Glutamate receptor response properties of nociceptive synapses on neurokinin 1 receptor positive (NK1R+) lamina I neurons were determined 3 days after induction of chronic peripheral inflammation with Freund's Complete Adjuvant (CFA). A significant increase in the AMPAR/NMDAR ratio was found during inflammation, which was associated with a significant reduction in the quantal amplitude of NMDAR-mediated synaptic currents. A significant shortening of the quantal AMPA current decay, a greater inward rectification of the AMPAR-mediated eEPSC amplitude and an increased sensitivity to the Ca2+-permeable AMPAR channel blocker 1-naphthylacetyl spermine (NAS) was also observed, indicating an increase in the contribution of Ca2+-permeable AMPARs at this synapse during inflammation. Furthermore the reduced effectiveness of the NR2B-specific antagonist CP-101,606 on NMDAR-mediated eEPSCs together with a decrease in Mg2+ sensitivity suggests a down regulation of the highly Mg2+-sensitive and high conductance NR2B subunit at this synapse. These changes in glutamatergic receptor function during inflammation support the selective effectiveness of Ca2+-permeable AMPAR antagonists in inflammatory pain models and may underlie the reported ineffectiveness of NR2B antagonists in spinal antinociception.
Asunto(s)
Calcio/metabolismo , Conducción Nerviosa/fisiología , Dolor/metabolismo , Células del Asta Posterior/metabolismo , Receptores AMPA/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , Sinapsis/metabolismo , Animales , Potenciales Postsinápticos Excitadores/fisiología , Femenino , Adyuvante de Freund , Inflamación/inducido químicamente , Inflamación/complicaciones , Inflamación/metabolismo , Masculino , Dolor/etiología , Ratas , Ratas Sprague-Dawley , Receptores de Neuroquinina-1/metabolismo , Receptores Opioides/metabolismo , Receptor de NociceptinaRESUMEN
The proinflammatory cytokine interferon-gamma (IFN-gamma), which can be present in elevated levels in the central nervous system during pathological conditions, may be involved in the generation of persistent pain states by inducing neuronal hyperexcitability. The aim of the present study was to examine whether loss of dorsal horn GABAergic inhibition may underlie this IFN-gamma-mediated neuronal hyperexcitability. Repetitive intrathecal injections of recombinant rat IFN-gamma (1000 U) or control buffer were administered to rats every second day for eight days. Electrophysiological recordings from lumbar dorsal horn neurons (n=46) were performed under halothane anaesthesia. Cellular responses were recorded before, during and after microiontophoretic application of the GABA antagonist bicuculline. In control animals, all cellular responses studied were significantly enhanced in the presence of bicuculline, including increased spontaneous activity, enhanced responses to innocuous and noxious mechanical stimulation and reduced paired-pulse depression. In contrast, in IFN-gamma-treated animals, bicuculline ejection had little or no facilitating effect on neuronal responses and instead a significant proportion of neurons displayed reduced responses. Seventy-four percent of cells from IFN-gamma treated animals showed a reduction in the response to noxious stimulation and 47% of the cells showed increased rather than reduced paired-pulse depression in the presence of bicuculline, thus suggesting IFN-gamma-induced excitatory actions by GABA. These findings show that the prolonged presence of increased levels of IFN-gamma in the central nervous system may contribute to the generation of central sensitization and persistent pain by reducing inhibitory tone in the dorsal horn. This implies a potential link between disinhibition and cytokine action in the spinal cord.
Asunto(s)
Interferón gamma/farmacología , Inhibición Neural/efectos de los fármacos , Células del Asta Posterior/efectos de los fármacos , Médula Espinal/citología , Ácido gamma-Aminobutírico/metabolismo , Potenciales de Acción/efectos de los fármacos , Análisis de Varianza , Animales , Bicuculina/farmacología , Distribución de Chi-Cuadrado , Interacciones Farmacológicas , Antagonistas del GABA/farmacología , Inyecciones Espinales/métodos , Masculino , Umbral del Dolor/efectos de los fármacos , Estimulación Física/métodos , Ratas , Ratas Wistar , Piel/inervaciónRESUMEN
N-type calcium channels contribute to the release of glutamate from primary afferent terminals synapsing onto nocisponsive neurons in the dorsal horn of the spinal cord, but little is known of functional adaptations to these channels in persistent pain states. Subtype-selective conotoxins and other drugs were used to determine the role of different calcium channel types in a rat model of inflammatory pain. Electrically evoked primary afferent synapses onto lumber dorsal horn neurons were examined three days after induction of inflammation with intraplantar complete Freund's adjuvant. The maximal inhibitory effect of the N-type calcium channel blockers, omega-conotoxins CVID and MVIIA, were attenuated in NK1 receptor-positive lamina I neurons after inflammation, but the potency of CVID was unchanged. This was associated with reduced inhibition of the frequency of asynchronous-evoked synaptic events by CVID studied in the presence of extracellular strontium, suggesting reduced N-type channel contribution to primary afferent synapses after inflammation. After application of CVID, the relative contributions of P/Q and L channels to primary afferent transmission and the residual current were unchanged by inflammation, suggesting the adaptation was specific to N-type channels. Blocking T-type channels did not affect synaptic amplitude under control or inflamed conditions. Reduction of N-type channel contribution to primary afferent transmission was selective for NK1 receptor-positive neurons identified by post hoc immunohistochemistry and did not occur at synapses in laminae II(o) or II(i), or inhibitory synapses. These results suggest that inflammation selectively downregulates N-type channels in the terminals of primary afferents synapsing onto (presumed) nociceptive lamina I NK1 receptor-positive neurons.
Asunto(s)
Canales de Calcio Tipo N/metabolismo , Inflamación/fisiopatología , Células del Asta Posterior/metabolismo , Receptores de Neuroquinina-1/metabolismo , Transmisión Sináptica , Vías Aferentes/fisiopatología , Animales , Canales de Calcio Tipo N/efectos de los fármacos , Enfermedad Crónica , Regulación hacia Abajo , Estimulación Eléctrica , Potenciales Evocados/efectos de los fármacos , Femenino , Adyuvante de Freund , Miembro Posterior , Inflamación/inducido químicamente , Inflamación/metabolismo , Masculino , Nociceptores , Dolor/metabolismo , Dolor/fisiopatología , Ratas , Ratas Sprague-Dawley , Médula Espinal/fisiopatología , Sinapsis , Ponzoñas/farmacología , omega-ConotoxinasRESUMEN
Hyperexcitability of spinal dorsal horn neurons, also known as 'central sensitization', is a component of pain associated with pathological conditions in the nervous system. The aim of the present study was to analyze if the pro-inflammatory cytokine, interferon-gamma (IFN-gamma), which can be released for extended periods of time in the nervous system during inflammatory and infectious events, can alter synaptic activity in dorsal horn neurons and thereby contribute to such hyperexcitability. Treatment of cultured dorsal horn neurons with IFN-gamma for 2 weeks resulted in a significantly reduced clustering of alpha-amino-3-hydroxy-5-methylisoxazole (AMPA) receptor subunit 1 (GluR1) that was dependent on nitric oxide. The neurons displayed an increased frequency and amplitude of excitatory postsynaptic currents (EPSCs) upon IFN-gamma treatment. Treated dorsal horn neurons also exhibited increased responsiveness to stimulation of dorsal root ganglia (DRG) axons in a two-compartment model. Furthermore, disinhibition by the GABA(A) receptor antagonist picrotoxin (PTX) significantly increased EPSC frequency and induced bursting in untreated cultures but did not significantly increase the frequency in treated neurons, which displayed bursting even without PTX. GABA(A) agonists reduced activity more strongly in treated cultures and immunochemical staining for GABA(A) receptors showed no difference from controls. Since GluR1-containing AMPA receptors (AMPARs) occur predominantly on inhibitory neurons in the dorsal horn, we suggest that the IFN-gamma-mediated increase in spontaneous activity and responsiveness to DRG axon stimulation, decrease in sensitivity to PTX and tendency for EPSC bursting result from a reduced expression of GluR1 on these neurons and not from a reduction in active GABA(A) receptors in the network. IFN-gamma thereby likely causes disinhibition of synaptic activity and primary afferent input in the dorsal horn, which consequently results in central sensitization.
Asunto(s)
Potenciales Postsinápticos Excitadores/efectos de los fármacos , Interferón gamma/farmacología , Células del Asta Posterior/efectos de los fármacos , Animales , Células Cultivadas , Embrión de Mamíferos , Potenciales Postsinápticos Excitadores/fisiología , Femenino , Células del Asta Posterior/fisiología , Embarazo , Ratas , Ratas Sprague-Dawley , Receptores AMPA/fisiologíaRESUMEN
Long-term potentiation (LTP) of transmission of impulses in unmyelinated (C-fibre) primary afferents by prior tetanic conditioning stimulation has been demonstrated in the dorsal horn of the spinal cord. Since this potentiation has been proposed to be relevant to the increased responsiveness of spinal neurones associated with peripheral inflammation (central sensitisation), the present experiments compared the induction of LTP in normal rats and rats with monoarthritis. Monoarthritis was induced by injection of complete Freund's adjuvant (CFA) into the left ankle joint of 12 rats. All animals showed behavioural signs of thermal hyperalgesia and were used for electrophysiological experiments after 4-8 days. In each animal, extracellular recordings were obtained from a single, wide dynamic range (WDR) dorsal horn neurone. High frequency tetanic conditioning stimulation of the sciatic nerve gave varying effects on the C-fibre-evoked responses of neurones in the normal rats, with potentiation in two, no change in five and a depression in five. By contrast, conditioning stimulation in rats with inflammation produced a long-lasting potentiation of C-fibre-evoked responses in 11 out of 12 neurones, with no effect in one. The ease with which LTP was induced in animals with inflammation supports the proposal that the underlying mechanisms of LTP are similar to those of the central sensitisation associated with peripheral inflammation.