RESUMO
These studies have demonstrated that ipsilateral renal artery occlusion (RAO) in rat results in the phosphorylation of cyclic AMP (cAMP) response element binding protein (p-CREB) in the thoracolumbar (T8-L2) spinal cord and associated dorsal root ganglia (DRG). p-CREB-immunoreactivity (IR) was expressed bilaterally in the thoracolumbar spinal cord, whereas expression in the DRG was ipsilateral relative to RAO. p-CREB-IR was primarily expressed in four distinct regions of the spinal cord: medial or lateral dorsal horn (MDH or LDH), dorsal commissural nucleus (DCN) and the region of the intermediolateral cell column (IML). After RAO, p-CREB-IR was greatest in the T13-L2 spinal segments. Within the T13-L1 spinal segments, p-CREB-IR was greatest in the MDH, LDH and DCN and expression in each of these regions was comparable within a segment. Following RAO, there was a significant (p < or = 0.001) increase in the percentage (86-98%) of p-CREB-IR spinal neurons expressing choline acetyltransferase (ChAT)-IR (a marker of preganglionic neurons) in the IML of the T10, T12 and L1 spinal segments examined. After ipsilateral RAO, expression of p-CREB-IR was increased in the ipsilateral, T8-L2 DRG with the greatest number of p-CREB-IR dorsal root ganglion cells being located in the L1 dorsal root ganglion. Retrograde tracing with Fluorogold (FG) to label renal afferent cells in the DRG revealed a significant (p < or = 0.01) increase in the percentage (75-86%) of renal afferent cells expressing p-CREB-IR after ipsilateral RAO. These studies demonstrate that p-CREB-IR is a useful tool for examining the distribution of spinal neurons and DRG involved in reflexes of renal origin. In addition, expression of p-CREB-IR may be coupled to late response genes that may exert long-term changes in neuronal function after RAO.
Assuntos
Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Gânglios Espinais/metabolismo , Neurônios/metabolismo , Artéria Renal/fisiologia , Medula Espinal/metabolismo , Animais , Fibras Autônomas Pré-Ganglionares/metabolismo , Colina O-Acetiltransferase/metabolismo , Feminino , Gânglios Espinais/citologia , Imuno-Histoquímica , Masculino , Óxido Nítrico Sintase/metabolismo , Óxido Nítrico Sintase Tipo I , Fosforilação , Ratos , Ratos Wistar , Medula Espinal/citologiaRESUMO
In cultured cerebrocortical neurons, mild excitotoxic insults or staurosporine result in apoptosis. We show here that N-methyl-d-aspartate (NMDA) receptor-mediated, but not staurosporine-mediated, apoptosis is preceded by depolarization of the mitochondrial membrane potential (Deltapsi(m)) and ATP loss. Both insults, however, release cytochrome c (Cyt c) into the cytoplasm. What prompts mitochondria to release Cyt c and the mechanism of release are as yet unknown. We examined the effect of inhibition of the adenine nucleotide translocator (ANT), a putative component of the mitochondrial permeability transition pore. Inhibition of the mitochondrial ANT with bongkrekic acid (BA) prevented NMDA receptor-mediated apoptosis of cerebrocortical neurons. Concomitantly, BA prevented Deltapsi(m) depolarization, promoted recovery of cellular ATP content, and blocked caspase-3 activation. However, in the presence of BA, Cyt c was still released. Because BA prevented NMDA-induced caspase-3 activation and apoptosis, the presence of Cyt c in the neuronal cytoplasm is not sufficient for the induction of caspase activity or apoptosis. In contrast to these findings, BA was ineffective in preventing staurosporine-induced activation of caspases or apoptosis. Additionally, staurosporine-induced, but not NMDA-induced, apoptosis was associated with activation of caspase-8. These results indicate that, in cerebrocortical cultures, excessive NMDA receptor activation precipitates neuronal apoptosis by means of mitochondrial dysfunction, whereas staurosporine utilizes a distinct pathway.