6-cyano-7-nitroquinoxaline-2-3-dione and Brain-Infarction

NMDA-type glutamate receptors mediate both trophic and excitotoxic signalling in CNS neurons. We have previously shown that blocking NMDAR- post-synaptic density-95 (PSD95) interactions provides significant protection from excitotoxicity and in vivo ischaemia; however, the mechanism of neuroprotection is unclear. Here, we report that blocking PSD-95 interactions with the Tat-NR2B9c peptide enhances a Ca²⁺-dependent protective pathway converging on cAMP Response Element binding protein (CREB) activation. We provide evidence that Tat-NR2B9c neuroprotection from oxygen glucose deprivation and NMDA toxicity occurs in parallel with the activation of calmodulin kinase signalling and is dependent on a sustained phosphorylation of the CREB transcription factor and its activator CaMKIV. Tat-NR2B9c-dependent neuroprotection and CREB phosphorylation are blocked by coapplication of CaM kinase (KN93 and STO-609) or CREB (KG-501) inhibitors, and by siRNA knockdown of CaMKIV. These results are mirrored in vivo in a rat model of permanent focal ischaemia. Tat-NR2B9c application significantly reduces infarct size and causes a selective and sustained elevation in CaMKIV phosphorylation; effects which are blocked by coadministration of KN93. Thus, calcium-dependent nuclear signalling via CaMKIV and CREB is critical for neuroprotection via NMDAR-PSD95 blockade, both in vitro and in vivo. This study highlights the importance of maintaining neuronal function following ischaemic injury. Future stroke research should target neurotrophic and pro-survival signal pathways in the development of novel neuroprotective strategies.

Topics: 6-Cyano-7-nitroquinoxaline-2,3-dione; Animals; Brain Infarction; Calcium Channel Blockers; Calcium-Calmodulin-Dependent Protein Kinase Type 4; Cells, Cultured; Cerebral Cortex; CREB-Binding Protein; Disease Models, Animal; Disks Large Homolog 4 Protein; Embryo, Mammalian; Enzyme Activation; Enzyme Inhibitors; Excitatory Amino Acid Antagonists; Glucose; Hypoxia; In Vitro Techniques; Infarction, Middle Cerebral Artery; Intracellular Signaling Peptides and Proteins; Male; Membrane Proteins; Neurons; Nimodipine; Phosphorylation; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Time Factors

Recent advances in functional imaging of human brain activity in stroke patients, e.g., functional magnetic resonance imaging, have revealed that cortical hemisphere contralateral to the infarction plays an important role in the recovery process. However, underlying mechanisms occurring in contralateral hemisphere during functional recovery have not been elucidated. We experimentally induced a complete infarction of somatosensory cortex in right hemisphere of mice and examined the neuronal changes in contralateral (left) somatosensory cortex during recovery. Both basal and ipsilateral somatosensory stimuli-evoked neuronal activity in left (intact) hemisphere transiently increased 2 d after stroke, followed by an increase in the turnover rate of usually stable mushroom-type synaptic spines at 1 week, observed by using two-photon imaging in vivo. At 4 weeks after stroke, when functional recovery had occurred, a new pattern of electrical circuit activity in response to somatosensory stimuli was established in intact ipsilateral hemisphere. Thus, the left somatosensory cortex can compensate for the loss of the right somatosensory cortex by remodeling neuronal circuits and establishing new sensory processing. This finding could contribute to establish the effective clinical treatments targeted on the intact hemisphere for the recovery of impaired functions and to achieve better quality of life of patients.

Topics: 6-Cyano-7-nitroquinoxaline-2,3-dione; Action Potentials; Animals; Brain Infarction; Dendritic Spines; Excitatory Amino Acid Antagonists; Functional Laterality; Luminescent Proteins; Male; Mice; Mice, Inbred C57BL; Mice, Transgenic; Neuronal Plasticity; Neurons; Physical Stimulation; Recovery of Function; Somatosensory Cortex; Stroke; Synapses; Time Factors