RESUMEN
BACKGROUND: Previous studies found that visceral sensitivity is increased in bowel obstruction (BO). We hypothesized that mechanical stress-induced expression of BDNF in smooth muscle cells (SMC) of the distended bowel plays a critical role in visceral hypersensitivity in BO by altering voltage-gated K+ channel (Kv ) activity in sensory neurons. METHODS: Partial colon obstruction was maintained in rats for 7 days. Colon-projecting neurons in the dorsal root ganglia (DRG, T13 to L2) were isolated for electrophysiological and gene expression studies. KEY RESULTS: Compared to controls, membrane excitability of colon-projecting DRG neurons was markedly enhanced in BO. The densities of total Kv and transient A-type (IA ) K+ currents, but not sustained delayed IK current, were significantly reduced in the neurons in BO. The mRNA expression of IA subtype Kv 1.4 in colon neurons was down-regulated in BO. Expression of BDNF mRNA and protein was dramatically increased in colonic smooth muscle of the distended segment, but not in the non-distended aboral segment. Mechanical stretch of colon SMC in vitro increased BDNF expression. Treatment with anti-BDNF antibody restored total Kv and IA currents of neurons from BO rats. Administration of Trk B inhibitor ANA-12 blocked BO-associated changes of neuronal excitability, Kv activity and gene expression in obstruction. CONCLUSIONS AND INFERENCES: Mechanical stress-induced expression of BDNF in colon SMC plays a critical role in visceral hypersensitivity in BO by suppressing A-type K+ currents and gene expression in sensory nerve. These findings help to identify therapeutic targets for distention-associated abdominal pain in the gut.
Asunto(s)
Dolor Abdominal/metabolismo , Factor Neurotrófico Derivado del Encéfalo/metabolismo , Ganglios Espinales/metabolismo , Hiperalgesia/metabolismo , Obstrucción Intestinal/metabolismo , Dolor Abdominal/etiología , Animales , Obstrucción Intestinal/complicaciones , Masculino , Ratas , Ratas Sprague-Dawley , Células Receptoras Sensoriales/metabolismo , Estrés MecánicoRESUMEN
It has been generally assumed that the cell body (soma) of a neuron, which contains the nucleus, is mainly responsible for synthesis of macromolecules and has a limited role in cell-to-cell communication. Using sniffer patch recordings, we show here that electrical stimulation of dorsal root ganglion (DRG) neurons elicits robust vesicular ATP release from their somata. The rate of release events increases with the frequency of nerve stimulation; external Ca(2+) entry is required for the release. FM1-43 photoconversion analysis further reveals that small clear vesicles participate in exocytosis. In addition, the released ATP activates P2X7 receptors in satellite cells that enwrap each DRG neuron and triggers the communication between neuronal somata and glial cells. Blocking L-type Ca(2+) channels completely eliminates the neuron-glia communication. We further show that activation of P2X7 receptors can lead to the release of tumor necrosis factor-alpha (TNFalpha) from satellite cells. TNFalpha in turn potentiates the P2X3 receptor-mediated responses and increases the excitability of DRG neurons. This study provides strong evidence that somata of DRG neurons actively release transmitters and play a crucial role in bidirectional communication between neurons and surrounding satellite glial cells. These results also suggest that, contrary to the conventional view, neuronal somata have a significant role in cell-cell signaling.
Asunto(s)
Adenosina Trifosfato/metabolismo , Comunicación Celular/fisiología , Ganglios Espinales/metabolismo , Neuronas/metabolismo , Células Satélites Perineuronales/metabolismo , Análisis de Varianza , Calcio/metabolismo , Línea Celular , Estimulación Eléctrica , Exocitosis/fisiología , Ganglios Espinales/fisiología , Humanos , Neuronas/fisiología , Compuestos de Piridinio , Compuestos de Amonio Cuaternario , Receptores Purinérgicos P2/metabolismo , Receptores Purinérgicos P2X7 , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
To develop a genetic approach for the treatment of pain, we introduced a recombinant adeno-associated viral (rAAV) vector containing the cDNA for the mu-opioid receptor (muOR) into primary afferent neurons in dorsal root ganglia (DRGs) of rats, which resulted in a long-lasting (>6 months) increase in muOR expression in DRG neurons. The increase greatly potentiated the antinociceptive effects of morphine in rAAV-muOR-infected rats with and without inflammation. Perforated patch recordings indicated that the efficacy and potency of opioid inhibition of voltage-dependent Ca(2+) channels were enhanced in infected neurons, which may underlie the increase in opiate efficacy. These data suggest that transfer of opioid receptor genes into DRG cells with rAAV vectors may offer a new therapeutic strategy for pain management.
Asunto(s)
Dependovirus/genética , Morfina/farmacología , Neuronas Aferentes/fisiología , Dolor/prevención & control , Receptores Opioides/genética , Animales , Ganglios Espinales/fisiología , Ganglios Espinales/fisiopatología , Técnicas de Transferencia de Gen , Proteínas Fluorescentes Verdes , Proteínas Luminiscentes/genética , Potenciales de la Membrana/fisiología , Modelos Neurológicos , Neuronas Aferentes/efectos de los fármacos , Técnicas de Placa-Clamp , Plásmidos , Ratas , Ratas Sprague-Dawley , Receptores Opioides/fisiología , Proteínas Recombinantes de Fusión/metabolismo , Proteínas Recombinantes/metabolismoRESUMEN
We previously reported that gabapentin (GBP), a widely prescribed analgesic, enhances N-methyl-aspartate (NMDA) receptor mediated currents only when the intracellular level of protein kinase C is elevated. However, it is unclear how the potentiation of NMDA responses by GBP can lead to pain relief. To resolve this issue, we combined immunocytochemical and patch recording techniques to study the actions of GBP on NMDA receptors in dorsal horn cells isolated from rats with inflammation and to determine the gamma-aminobutyric acid (GABA) content in the recorded cells. We found that all GBP-responsive cells are GABA-immunoreactive and none of the GABA-negative neurons respond to GBP. Thus, GBP appears to enhance NMDA currents in GABAergic neurons. These observations suggest that GBP exerts its antinociceptive action by increasing the activity of these inhibitory neurons.