Inhibition of Nitric Oxide Production and Protein Tyrosine Nitration Contribute to Neuroprotection by a Novel Calmodulin Antagonist, DY-9760e, in the Rat Microsphere Embolism

Abstract

Human multiple small cerebral infarcts are implicated in induction of stroke attacks and dementia associated with multiple brain embolism. These disorders were caused by permanent or prolonged occlusion of multiple microvascular in the brain. Unfortunately, few animal models can mimic clinical features in human multiple brain embolism. Recent evidences, however, suggest that microsphere embolism (ME)-induced ischemia model in rat resembles to multiple embolism or multi-infarct dementia in human with the clinical features.2,3) The cerebral multiple embolism is performed by injection of non-radioactive microsphere with a diameter ranging from 15 to 50 mm into the unilateral internal carotid artery in the rat. The light microscopic analysis or 2,3,5-triphenyltetrazolium chloride (TTC) staining shows widespread scattered infarct in the cerebral cortex, striatum and hippocampus of the ipsilateral hemisphere injected microsphere. The intra-arterial infusion of microsphere also leads blood-brain barrier disruption and cerebral vascular injury, thereby inducing severe brain edema. 4,5) Furthermore, during and after ME-induced ischemia, abnormal intracellular signal transduction underlying detrimental metabolic and morphologic neuronal damages was observed.6) However, little neurochemical events through nitric oxide (NO) production are known in the rodent multiple embolism model.During cerebral ischemia, an abnormal elevation of intracellular Ca 2ϩ triggers activation of various Ca 2ϩ -dependent enzymes including endonucleases, proteases and phospholipases, thereby leading neuronal cell death.7-10) Among Ca 2ϩ -mediated cell death pathways, calmodulin (CaM) is partly involved in the detrimental events leading neuronal injury. [11][12][13] In fact, CaM antagonists such as W-7 and calmidazolium have neuroprotective effects on NMDA-induced neuronal cell death 14) and trifluoperazine reduced infarct volume in transient focal cerebral ischemia.15) Similarly, FK-506, an immunosuppressant inhibiting calcineurin, exerts neuroprotective actions in glutamate neurotoxity.16) Since neuronal nitric oxide synthase (nNOS) knockout mice showed a reduced infarct size following brain ischemia, nNOS underlies detrimental events in the brain ischemia. 17)We recently introduced a novel calmodulin antagonist, DY9760e, having a potent inhibitory effect on Ca 2ϩ /calmodulindependent nNOS.18,19) DY-9760e rescues neurons from the ischemia/reperfusion-induced neuronal death with concomitant inhibition of NO production during ischemia/reperfusion in the gerbil hippocampus.20) Ischemia/reperfusion-induced neuronal injury is also triggered by peroxynitrite (ONOO Ϫ ), which is formed by the rapid reaction of NO with superoxide (O 2 Ϫ ).21) The protein tyrosine nitration during ischemia/ reperfusion was also prevented by DY-9760e treatment in the gerbil ischemic brain. 20) Thus, the detrimental effects of NO and protein tyrosine nitration were defined in the ischemia/ reperfusion-induced neuronal injury. However, the detrimental effects of NO productio...

Postischemic hyperoxia reduces hippocampal pyruvate dehydrogenase activity

“…4-6). Tyrosine nitration is involved in many postischemic modifications [55][56][57]. Examples of proteins susceptible to inactivation by tyrosine nitration include mitochondrial superoxide dismutase [58,59] and several enzymes involved in oxidative energy metabolism, including aconitase [60], glutamate dehydrogenase [46], α-ketoglutarate dehydrogenase [61], and PDHC [46].…”

Antioxidant, antiinflammatory and antiapoptotic effects of dapsone in a model of brain ischemia/reperfusion in rats

“…In addition, during the ischemic period, ATP depletion causes a decrease in sodium-potassium ATPase pump activity, which in turn generates a disruption in the plasma membrane potential, leading to release of glutamate and other neurotransmitters (Globus et al, 1988;Santos et al, 1996;Melani et al, 1999). This anomalous increase in extracellular excitatory neurotransmitters produces excitotoxicity, accompanied by a rise in the cytosolic calcium concentrations, in turn associated with activation of calcium-dependent enzymes, such as calpain, calcineurin, and phospholipases, with a subsequent proteolysis of calpain substrates, activation of constitutive nitric oxide synthase (cNOS) enzyme, and release of arachidonic acid (White et al, 2000;Pisani et al, 2004;Shirakura et al, 2005). These processes are particularly important after ischemia insofar as they initiate a cascade of inflammatory responses, overproduction of free radicals, and activation of apoptosis (Zheng and Yenari, 2004;Chan, 2004;Margaill et al, 2005).…”

3-[2-[4-(3-Chloro-2-methylphenylmethyl)-1-piperazinyl]ethyl]-5,6-dimethoxy-1-(4-imidazolylmethyl)-1H-indazole Dihydro-chloride 3.5 Hydrate (DY-9760e) Is Neuroprotective in Rat Microsphere Embolism: Role of the Cross-Talk between Calpain and Caspase-3 through Calpastatin

“…Brain ischemia/reperfusion injury is associated with calpain-induced proteolysis of the cytoskeleton, including fodrin/spectrin (Fokuda et al, 1998;Sato et al, 1999;Yakota et al, 2003). We also documented nitric oxide production through activation of Ca 2ϩ /CaM-dependent nitric-oxide synthase (NOS) in ME-induced ischemia, thereby producing protein tyrosine nitration (Shirakura et al, 2005). The novel CaM inhibitor DY-9760e rescues neurons from ME-induced neuronal damage with concomitant inhibition of both NO production and protein tyrosine nitration (Hashiguchi et al, 2003).…”