Alterations Induced by Ischemic Preconditioning on Secretory Pathways Ca2+-ATPase (SPCA) Gene Expression and Oxidative Damage After Global Cerebral Ischemia/Reperfusion in Rats

Abstract

Ischemic preconditioning (IPC) represents the phenomenon of CNC adaptation, which results in increased tolerance of CNS to lethal ischemia. Brain ischemia/reperfusion (IRI) initiates a catastrophic cascade in which many subcellular organelles play an important role. The Golgi apparatus, which is a part of secretory pathways (SP), represents the Ca(2+) store and regulates secretion of proteins for growth/reorganization of neuronal circuit by secretory Ca(2+)ATPases (SPCA1). The purpose of this study is to evaluate the effect of IRI and preconditioning on SPCA1 gene expression and oxidative damage after 4-vessel occlusion for 15 min and after being exposed to different reperfusion periods. Rats were preconditioned by 5 min of sub-lethal ischemia and 2 days later, 15 min of lethal ischemia was induced. Our experiments conclusively showed IRI-induced depression of SPCA activity and lipo- and protein oxidation in rat hippocampal membranes. IRI also activates the induction of SPCA1 gene expression in later reperfusion periods. IPC partially suppresses lipo- and protein oxidation in hippocampal membranes and leads to partiall rovery of the ischemic-induced depression of SPCA activity. In addition, IPC initiates earlier cellular response to the injury by the significant elevation of mRNA expression to 142% comparing to 1 h of corresponding reperfusion and to 11% comparing to 24 h of corresponding reperfusion, respectively. Similar patterns were observed on the translational level by Western blot analysis. Our results indicate the specific SPCA1 expression pattern in ischemic hippocampus. It also shows that the SPCA expression and the post-translational changes induced by ischemia are modulated by the IPC. This might serve to understand the molecular mechanisms involved in the structural integrity and function of the SP after ischemic challenge. It also suggests that there is a correlation of SPCA function with the role of SP in the response to pre-ischemic challenge.

Mechanisms of Ischemic Induced Neuronal Death and Ischemic Tolerance

“…In order to evaluate whether the severe metabolic stress induced by IRI and/or IPC affects transcription of SPCA1 gene, the mRNA and protein levels of SPCA1 was analyzed (Lehotsky, 1999(Lehotsky, , 2002a(Lehotsky, , 2004. As shown by Pavlikova et al (2009), RT-PCR clearly detected, that hippocampal cells respond to the IRI by induction of mRNA level in reperfusion period with maximum at 3 h reperfusion (to 171% of control). Preconditioning (IPC) initiates earlier tissue response to the injury by the significant elevation of mRNA expression already at 1 h of reperfusion and the level of mRNA expression reached 142% comparing to 1 h ischemia, and to 164% comparing to control.…”

“…The authors showed that SPCA activity, similar to other P-type ATPases, is also subject to ischemic damage most likely due to free radicals action (Lehotsky et al, 2002b). In addition, oxidative alterations detected in mitochondria and microsomes after IRI in our experiments, may at least partially explain functional postischemic disturbances of neuronal ion transport mechanisms (Lipton 1999;Lehotsky et al, 2002a;Obrenovitch, 2008) and inhibition of global proteosynthesis (Burda et al, 2003), which are both implicated in neuronal cell damage and/or recovery from ischemic insult, IPC-induced reduction of lipoperoxidation products and protein oxidative changes (Racay et al, 2009;Pavlikova et al, 2009). These may all be probably due to upregulation of defence mechanisms (antioxidant enzymes) against oxidative stress in the preconditioning challenge (Danielisova et al, 2005;Gidday, 2006;Obrenovitch, 2008).…”

“…Similar patterns were observed on the translational level by Western blot analysis. Results of Pavlikova et al (2009) indicate the specific SPCA1 expression pattern in injured ischemic hippocampus and might serve to understand the molecular mechanisms involved in the structural integrity and function of the Golgi complex after ischemic challenge. They also suggest for the correlation of SPCA function with the role of SP in response to preischemic challenge.…”

“…The molecular mechanisms underlying ischemic tolerance are not yet fully understood yet. Therefore a series of papers (Urban et al, 2009;Pavlikova et al,2009) have focused attention at the mRNA and protein levels of the ER stress genes after ischemic/reperfusion damage (IRI) in naive and preconditioned groups of rats.…”

“…Oxidative burst lasting several minutes upon the onset of reperfusion is followed by dysregulation of antioxidant mechanism and moderate but persistently elevated production of oxygen radicals which might initiate cell death signaling pathways after cerebral ischemia and parallels with selective postischemic vulnerability of the brain (Valko et al, 2007;Shi and Liu, 2007). One of the main aims of the study of Pavlikova et al (2009) was to determine whether IRI and IPC would affect the physical and functional properties of hippocampal membrane vesicles including Golgi SP. Neuronal microsomes are vulnerable to physical and functional oxidative damage (Lehotsky et al 1999(Lehotsky et al , 2002aUrikova et al 2006).…”