Leptin-mediated cell survival signaling in hippocampal neurons mediated by JAK STAT3 and mitochondrial stabilization

Leptin plays a pivotal role in the regulation of energy homeostasis and metabolism, primarily by acting on neurons in the hypothalamus that control food intake. However, leptin receptors are more widely expressed in the brain suggesting additional, as yet unknown, functions of leptin. Here we show that both embryonic and adult hippocampal neurons express leptin receptors coupled to activation of STAT3 and phosphatidylinositol 3-kinase-Akt signaling pathways. Leptin protects hippocampal neurons against cell death induced by neurotrophic factor withdrawal and excitotoxic and oxidative insults. The neuroprotective effect of leptin is antagonized by the JAK2-STAT3 inhibitor AG-490, STAT3 decoy DNA, and phosphatidylinositol 3-kinase/Akt inhibitors but not by an inhibitor of MAPK. Leptin induces the production of manganese superoxide dismutase and the anti-apoptotic protein Bcl-xL, and stabilizes mitochondrial membrane potential and lessens mitochondrial oxidative stress. Leptin receptor-deficient mice (db/db mice) are more vulnerable to seizure-induced hippocampal damage, and intraventricular administration of leptin protects neurons against seizures. By enhancing mitochondrial resistance to apoptosis and excitotoxicity, our findings suggest that leptin signaling serves a neurotrophic function in the developing and adult hippocampus.     

Hypoxic Preconditioning Attenuates Neuronal Cell Death by Preventing MEK/ERK Signaling Pathway Activation after Transient Global Cerebral Ischemia in Adult Rats

Our previous data indicated that hypoxic preconditioning (HPC) ameliorates transient global cerebral ischemia (tGCI)-induced neuronal death in hippocampal CA1 subregion of adult rats. However, the possible molecular mechanisms for neuroprotection of this kind are largely unknown. This study was performed to investigate the role of the mitogen-activated protein kinase/extra-cellular signal-regulated kinase kinase (MEK)/extra-cellular signal-regulated kinase (ERK) pathway in HPC-induced neuroprotection. tGCI was induced by applying the four-vessel occlusion method. Pretreatment with 30 min of hypoxia applied 1 day before 10 min tGCI significantly decreased the level of MEK1/2 and ERK1/2 phosphorylation in ischemic hippocampal CA1 subregion. Also, HPC decreased the expression of phosphorylated ERK1/2 in degenerating neurons and astrocytes. However, the administration of U0126, a MEK kinase inhibitor, partly blocked MEK1/2 and ERK1/2 phosphorylation induced by tGCI. Meanwhile, neuronal survival was improved, and glial cell activation was significantly reduced. Collectively, these data indicated that the MEK/ERK signaling pathway might be involved in HPC-induced neuroprotection following tGCI. Also, HPC resulted in a reduction of glial activation.     

NMDA and Non-NMDA Receptor Gene Expression Following Global Brain Ischemia in Rats: Effect of NMDA and Non-NMDA Receptor Antagonists

Abstract: Transient forebrain or global ischemia in rats induces selective and delayed damage of hippocampal CA1 neurons. In a previous sludy, we have shown that expression of GIuR2, the kainate/a-amino-3-hydroxy-5- methyl-4-isoxazolepropionic acid (AMPA) receptor subunit that governs Ca' permeability, is preferentially reduced in CA1 at a time point proceeding neuronal degeneration. Postischemic administration of the selective AMPA receptor antagonist, 2,3-dihydroxy-6-nitro-7-sulfamoylbenzo(f)quinoxaline (NBQX), protects CAI neurons against delayed death. In this study we examined the effects of NBQX (at a neuroprotective dose) and of MK-801 (a selective NMDA receptor anltagonist, not protective in this model) on kainate/AMPA receptor gene expression changes after global ischemia. We also examined the effects of transient forebrain ischemia on expression of the NMDA receptor subunit NMDARI. In ischemic rats treated with saline, GIuR2 and (31uR3 mRNAs were markedly reduced in CAI but were unchanged in CA3 or dentate gyrus. GluRl and NMDAR1 mRNAs were not significantly changed in any region examined. Administration of NBQX or MK-801 did not alter the ischemia-induced changes in kainate/AMPA receptor gene expression. These findings suggest that NBQX affords neuroprotection by a direct blockade of kainate/AMPA receptors, rather than by a modificatian of GIuR2 expression changes     

ERK 1/2 Activation Mediates the Neuroprotective Effect of BpV(pic) in Focal Cerebral Ischemia–Reperfusion Injury

Bisperoxovanadium (pyridine-2-carboxyl) [bpV(pic)] is a commercially available PTEN inhibitor. Previous studies from us and others have shown that bpV(pic) confers neuroprotection in cerebral ischemia injury. We set up to determine whether ERK 1/2 activation plays a role in bpV(pic)-induced neuroprotective effect in cerebral ischemia injury. We found that the phosphorylation levels of Akt (p-AKT) and ERK1/2 (p-ERK 1/2) were down-regulated after cerebral ischemia–reperfusion injury. The injection of bpV(pic) after injury not only increased the level of p-AKT but also the level of p-ERK 1/2. While the inhibition of PTEN mediated the up-regulatation of p-AKT and p-ERK 1/2 by bpV(pic). Interestingly, the ERK 1/2 activation induced by bpV(pic) was also independent of the inhibition of PTEN. Our results indicate that bpV(pic) protects against OGD-induced neuronal death and promotes the functional recovery of stroke animals through PTEN inhibition and ERK 1/2 activation, respectively. This study suggests that the effect of bpV(pic) on ERK 1/2 signaling should be considered while using bpV(pic) as a PTEN inhibitor.     

Insulin/PI3K signaling protects dentate neurons from oxygen–glucose deprivation in organotypic slice cultures

It is known that ischemia/reperfusion induces neurodegeneration in the hippocampus in a subregion‐dependent manner. This study investigated the mechanism of selective resistance/vulnerability to oxygen–glucose deprivation (OGD) using mouse organotypic hippocampal cultures. Analysis of propidium iodide uptake showed that OGD‐induced duration‐ and subregion‐dependent neuronal injury. When compared with the CA1–3 subregions, dentate neuronal survival was more sensitive to inhibition of phosphatidylinositol 3‐kinase (PI3K)/Akt signaling under basal conditions. Dentate neuronal sensitivity to PI3K/Akt signaling activation was inversely related to its vulnerability to OGD‐induced injury; insulin/insulin‐like growth factor 1 pre‐treatment conferred neuroprotection to dentate neurons via activation of PI3K/Akt signaling. In contrast, CA1 and CA3 neurons were less sensitive to disruptions of endogenous PI3K/Akt signaling and protective effects of insulin/insulin‐like growth factor 1, but more vulnerable to OGD. OGD‐induced injury in CA1 was reduced by inhibition of NMDA receptor or mitogen‐activated protein kinase signaling, and was prevented by blocking NMDA receptor in the presence of insulin. The CA2 subregion was distinctive in its response to glutamate, OGD, and insulin, compared with other CA subregions. CA2 neurons were sensitive to the protective effects of insulin against OGD‐induced injury, but more resistant to glutamate. Distinctive distribution of insulin receptor β and basal phospho‐Akt was detected in our slice cultures. Our results suggest a role for insulin signaling in subregional resistance/vulnerability to cerebral ischemia.     

Block of P2X7 receptors could partly reverse the delayed neuronal death in area CA1 of the hippocampus after transient global cerebral ischemia

Transient global ischemia (which closely resembles clinical situations such as cardiac arrest, near drowning or severe systemic hypotension during surgical procedures), often induces delayed neuronal death in the brain, especially in the hippocampal CA1 region. The mechanism of ischemia/reperfusion (I/R) injury is not fully understood. In this study, we have shown that the P2X7 receptor antagonist, BBG, reduced delayed neuronal death in the hippocampal CA1 region after I/R injury; P2X7 receptor expression levels increased before delayed neuronal death after I/R injury; inhibition of the P2X7 receptor reduced I/R-induced microglial microvesicle-like components, IL-1β expression, P38 phosphorylation, and glial activation in hippocampal CA1 region after I/R injury. These results indicate that antagonism of the P2X7 receptor and signaling pathways of microglial MV shedding, such as src-protein tyrosine kinase, P38 MAP kinase and A-SMase, might be a promising therapeutic strategy for clinical treatment of transient global cerebral I/R injury.     

Critical role of PTEN in the coupling between PI3K/Akt and JNK1/2 signaling in ischemic brain injury

JNK pathway is an important pro-apoptotic kinase cascade mediating cell death in response to a variety of extracellular stimuli including excitotoxicity, which results in selective and delayed neuronal death in the hippocampal CA1. On the contrary, activation of the protein kinase Akt, which is controlled by the opposing actions of PI3K and PTEN, contributes to enhanced resistance to apoptosis through multiple mechanisms. We here demonstrate that the temporal pattern of Akt activation reversely correlates with JNK1/2 activation following various time points of ischemic reperfusion. However, the activation of JNK1/2 could be decreased by the elevation of Akt activation via increasing the tyrosine phosphorylation of PTEN by bpv(pic), a potent PTPases inhibitor for PTEN, or by intracerebroventricular infusion of PTEN antisense oligodeoxynucleotides (AS-ODNs). In contrast, JNK1/2 activation was significantly increased by preventing PTEN degradation after pretreatment with proteasome inhibitor. The neuroprotective effects of bpv(pic) and PTEN AS-ODNs were significant in the CA1 subfield after transient global ischemia. In conclusion, the present results clearly show that PTEN plays a key regulatory role in the cross-talk between cell survival PI3K/Akt pathway and pro-death JNK pathway, and raise a new possibility that agents targeting phosphatase PTEN may offer a great promise to expand the therapeutic options in protecting neurons form ischemic brain damage.     

K252a suppresses neuronal cells apoptosis through inhibiting the translocation of Bax to mitochondria induced by the MLK3/JNK signaling after transient global brain ischemia in rat hippocampal CA1 subregion

It is demonstrated that the c-Jun N-terminal kinase (JNK) signaling pathway plays a critical role in ischemic brain injury. Our previous studies have suggested that K252a can obviously inhibit JNK activation induced by ischemia/reperfusion in the vulnerable hippocampal CA1 subregion. Here, we further discussed the potential mechanism of ischemic brain injury induced by the activation of JNK after 15?min of transient global cerebral ischemia. As a result, through inhibiting phosphorylation of Bcl-2 (a cytosolic target of JNK) and 14-3-3 protein (a cytoplasmic anchor of Bax) induced by the activation of JNK, K252a decreased the release of Bax from Bcl-2/Bax and 14-3-3/Bax dimers, further attenuating the translocation of Bax from cytosol to mitochondria and the release of cytochrome c induced by ischemia/reperfusion, which related to mitochondria-mediated apoptosis. Importantly, pre-infusion of K2525a 20?min before ischemia showed neuroprotective effect against neuronal cells apoptosis. These findings imply that K252a induced neuroprotection against ischemia/reperfusion in rat hippocampal CA1 subregion via inhibiting the mitochondrial apoptosis pathway induced by JNK activation.     

Estradiol rescues neurons from global ischemia-induced cell death: Multiple cellular pathways of neuroprotection

The potential neuroprotective role of sex hormones in chronic neurodegenerative disorders and acute brain ischemia following cardiac arrest and stroke is of a great therapeutic interest. Long-term pretreatment with estradiol and other estrogens affords robust neuroprotection in male and female rodents subjected to focal and global ischemia. However, the receptors (e.g., cell surface or nuclear), intracellular signaling pathways and networks of estrogen-regulated genes that intervene in neuronal apoptosis are as yet unclear. We have shown that estradiol administered at physiological levels for two weeks before ischemia rescues neurons destined to die in the hippocampal CA1 and ameliorates ischemia-induced cognitive deficits in ovariectomized female rats. This regimen of estradiol treatment involves classical intracellular estrogen receptors, transactivation of IGF-1 receptors and stimulation of the ERK/MAPK signaling pathway, which in turn maintains CREB activity in the ischemic CA1. We also find that a single, acute injection of estradiol administrated into the brain ventricle immediately after an ischemic event reduces both neuronal death and cognitive deficits. Because these findings suggest that hormones could be used to treat patients when given after brain ischemia, it is critical to determine whether the same or different pathways mediate this form of neuroprotection. We find that an agonist of the membrane estrogen receptor GPR30 mimics short latency estradiol facilitation of synaptic transmission in the hippocampus. Therefore, we are testing the hypothesis that GPR30 may act together with intracellular estrogen receptors to activate cell signaling pathways to promote neuron survival after global ischemia.     

Atorvastatin Attenuates Ischemia/Reperfusion-Induced Hippocampal Neurons Injury Via Akt-nNOS-JNK Signaling Pathway

Ischemia-induced brain damage leads to apoptosis like delayed neuronal death in selectively vulnerable regions, which could further result in irreversible damages. Previous studies have demonstrated that neurons in the CA1 area of hippocampus are particularly sensitive to ischemic damage. Atorvastatin (ATV) has been reported to attenuate cognitive deficits after stroke, but precise mechanism for neuroprotection remains unknown. Therefore, the aims of this study were to investigate the neuroprotective mechanisms of ATV against ischemic brain injury induced by cerebral ischemia reperfusion. In this study, four-vessel occlusion model was established in rats with cerebral ischemia. Rats were divided into five groups: sham group, I/R group, I/R+ATV group, I/R+ATV+LY, and I/R+SP600125 group. Cresyl violet staining was carried out to examine the neuronal death of hippocampal CA1 region. Immunoblotting was used to detect the expression of the related proteins. Results showed that ATV significantly protected hippocampal CA1 pyramidal neurons against cerebral I/R. ATV could increase the phosphorylation of protein kinase B (Akt1) and nNOS, diminished the phosphorylation of JNK3 and c-Jun, and further inhibited the activation of caspase-3. Whereas, all of the aforementioned effects of ATV were reversed by LY294002 (an inhibitor of Akt1). Furthermore, pretreatment with SP600125 (an inhibitor of JNK) diminished the phosphorylation of JNK3 and c-Jun, and further inhibited the activation of caspase-3 after cerebral I/R. Taken together, our results implied that Akt-mediated phosphorylation of nNOS is involved in the neuroprotection of ATV against ischemic brain injury via suppressing JNK3 signaling pathway that provide a new experimental foundation for stroke therapy.     

Increases of Catalase and Glutathione Peroxidase Expressions by Lacosamide Pretreatment Contributes to Neuroprotection Against Experimentally Induced Transient Cerebral Ischemia

Lacosamide is a new antiepileptic drug which is widely used to treat partial-onset seizures. In this study, we examined the neuroprotective effect of lacosamide against transient ischemic damage and expressions of antioxidant enzymes such as Zn-superoxide dismutase (SOD1), Mn-superoxide dismutase (SOD2), catalase (CAT) and glutathione peroxidase (GPX) in the hippocampal cornu ammonis 1 (CA1) region following 5 min of transient global cerebral ischemia in gerbils. We found that pre-treatment with 25 mg/kg lacosamide protected CA1 pyramidal neurons from transient global cerebral ischemic insult using hematoxylin–eosin staining and neuronal nuclear antigen immunohistochemistry. Transient ischemia dramatically changed expressions of SOD1, SOD2 and GPX, not CAT, in the CA1 pyramidal neurons. Lacosamide pre-treatment increased expressions of CAT and GPX, not SOD1 and 2, in the CA1 pyramidal neurons compared with controls, and their expressions induced by lacosamide pre-treatment were maintained after transient cerebral ischemia. In brief, pre-treatment with lacosamide protected hippocampal CA1 pyramidal neurons from ischemic damage induced by transient global cerebral ischemia, and the lacosamide-mediated neuroprotection may be closely related to increases of CAT and GPX expressions by lacosamide pre-treatment.     

Neuroprotective effects of preconditioning ischemia on ischemic brain injury through down-regulating activation of JNK1/2 via N-methyl-D-aspartate receptor-mediated Akt1 activation

Our previous studies have demonstrated that the JNK signaling pathway plays an important role in ischemic brain injury and is mediated via glutamate receptor 6. Others studies have shown that N-methyl-d-aspartate (NMDA) receptor is involved in the neuroprotection of ischemic preconditioning. Here we examined whether ischemic preconditioning down-regulates activation of the mixed lineage kinase-JNK signaling pathway via NMDA receptor-mediated Akt1 activation. In our present results, ischemic preconditioning could not only inhibit activations of mixed lineage kinase 3, JNK1/2, and c-Jun but also enhanced activation of Akt1. In addition, both NMDA (an agonist of NMDA receptor) and preconditioning showed neuroprotective effects. In contrast, ketamine, an antagonist of NMDA receptor, prevented the above effects of preconditioning. Further studies indicated that LY294002, an inhibitor of phosphoinositide 3-kinase that is an upstream signaling protein of Akt1, could block neuroprotection of preconditioning, and KN62, an inhibitor of calmodulin-dependent protein kinase, also achieved the same effects as LY294002. Therefore, both phosphoinositide 3-kinase and calmodulin-dependent protein kinase are involved in the activation of Akt1 in ischemic tolerance. Taken together, our results indicate that preconditioning can inhibit activation of JNK signaling pathway via NMDA receptor-mediated Akt1 activation and induce neuroprotection in hippocampal CA1 region.     

Ischemia-Induced Changes of PRAS40 and p-PRAS40 Immunoreactivities in the Gerbil Hippocampal CA1 Region After Transient Cerebral Ischemia

Proline-rich Akt substrate of 40-kDa (PRAS40) is one of the important interactive linkers between Akt and mTOR signaling pathways. The increase of PRAS40 is related with the reduction of brain damage induced by cerebral ischemia. In the present study, we investigated time-dependent changes in PRAS40 and phospho-PRAS40 (p-PRAS40) immunoreactivities in the hippocampal CA1 region of the gerbil after 5 min of transient cerebral ischemia. PRAS40 immunoreactivity in the CA1 region was decreased in pyramidal neurons from 12 h after ischemic insult in a time-dependent manner, and, at 5 days post-ischemia, PRAS40 immunoreactivity was newly expressed in astrocytes. p-PRAS40 immunoreactivity in the CA1 pyramidal neurons was hardly found 12 h and apparently detected again 1 and 2 days after ischemic insult. At 5 days post-ischemia, p-PRAS40 immunoreactivity in the CA1 pyramidal neurons was not found. These results indicate that ischemia-induced changes in PRAS40 and p-PRAS40 immunoreactivities in CA1 pyramidal neurons and astrocytes may be closely associated with delayed neuronal death in the hippocampal CA1 region following transient cerebral ischemia.     

Inhibition of the Connexin 43 Elevation May be Involved in the Neuroprotective Activity of Leptin Against Brain Ischemic Injury

Leptin is a multifunctional hormone produced by the ob gene and is secreted by adipocytes that regulate food intake and energy metabolism. Numerous studies demonstrated that leptin is a novel neuroprotective effector, however, the mechanisms are largely unknown. Herein, we demonstrate the protective activities of leptin after ischemic stroke and provide the first evidence for the involvement of the connexin 43 (Cx43) in leptin-mediated neuroprotection. We found that leptin treatment reduces the infarct volume, improves animal behavioral parameters, and inhibits the elevation of Cx43 expression in vivo. In vitro, leptin reverses ischemia-induced SY5Y and U87 cells Cx43 elevation, secreted glutamate levels in medium and SY5Y cell death, these roles could be abolished by leptin receptor blocker. Additionally, leptin administration upregulated the extracellular signal-regulated kinase1/2 (ERK1/2) phosphorylation. Moreover, ERK1/2 inhibitors pretreatment reversed the effects of leptin on Cx43 expression, glutamate levels and cell apoptosis. In conclusion, the present study demonstrated that leptin can reduce the Cx43 expression and cell death both in vivo and in vitro via ERK1/2 signaling pathway. This result provides a novel regulatory signaling pathway of the neuroprotective effects of leptin and may contribute to ischemic brain injury prevention and therapy.     

The delta-opioid receptor agonist DADLE at reperfusion protects the heart through activation of pro-survival kinases via EGF receptor transactivation

The specific delta-opioid receptor agonist [D-Ala(2)-D-Leu(5)]enkephalin (DADLE) protects against infarction in the heart when given before ischemia. In rabbit, this protection leads to phosphorylation of the pro-survival kinases Akt and extracellular signal-regulated kinase (ERK) and is dependent on transactivation of the epidermal growth factor receptor (EGFR). DADLE reportedly protects rat hearts at reperfusion. We therefore tested whether DADLE at reperfusion could protect isolated rabbit hearts subjected to 30 min of regional ischemia and 120 min of reperfusion and whether this protection is dependent on Akt, ERK, and EGFR. DADLE (40 nM) was infused for 1 h starting 5 min before reperfusion and reduced infarct size from 31.0 +/- 2.3% in the control group to 14.6 +/- 1.6% (P = 0.01). This protection was abolished by cotreatment of the metalloproteinase inhibitor (MPI) and the EGFR inhibitor AG1478. In contrast, 20 nM DADLE, although known to be protective before ischemia, failed to protect. Western blotting revealed that DADLE's protection was correlated to increase in phosphorylation of the kinases Akt and ERK1 and -2 in reperfused hearts (2.5 +/- 0.5, 1.6 +/- 0.2, and 2.3 +/- 0.7-fold of baseline levels, P < 0.05 vs. control). The DADLE-dependent increases in Akt and ERK1/2 phosphorylation were abolished by either MPI or AG1478, confirming a signaling through the EGFR pathway. Additionally, DADLE treatment increased phosphorylation of EGFR (1.4 +/- 0.2-fold, P = 0.03 vs. control). Thus the delta-opioid agonist DADLE protects rabbit hearts at reperfusion through activation of the pro-survival kinases Akt and ERK and is dependent on the transactivation of the EGFR.     

Preconditioning-induced activation of ERK5 is dependent on moderate Ca2+ influx via NMDA receptors and contributes to ischemic tolerance in the hippocampal CA1 region of rats

Accumulating evidence implicates activation (phosphorylation) of mitogen-activated protein kinases (MAPK) during nonlethal ischemic preconditioning in the protection of hippocampal CA1 neuron against subsequent ischemic events. In this paper, we undertook to identify the role of extracellular signal regulated kinase (ERK) 5 in cerebral ischemic preconditioning (CIP). Three minutes of ischemia was induced as preconditioning stimulus. Three days later, 6 min of ischemia was induced. The levels of ERK5 protein expression and its activation were detected with or without the CIP in hippocampal CA1 and the dentate gyrus (DG) regions. Our results showed that ERK5 was activated selectively in hippocampal CA1 region with, but not without, the ischemic preconditioning. Notably, during the later phase of reperfusion, the rise in ERK5 activation was strong and persistent with a peak occurring at the third day. The activation peak was effectively prevented and ERK5 protein expression was significantly decreased by intracerebroventricular infusion of ERK5 antisense oligonucleotide (every 24 h for 3 days before the preconditioning), but not by sense oligonucleotide or vehicle. Subsequently, the CA1 neuronal loss was largely elevated. Moreover, both MK801 (10 microM), an antagonist of NMDA receptor, and EGTA (100 mM, but neither 50 nor 150 mM), an extracellular Ca2+ chelator, not only effectively inhibited the ERK5 activation but also markedly abolished CIP-induced survival of the CA1 neurons. These results suggested that activation of the ERK5 pathway by CIP was at least partly dependent on moderate Ca2+ influx via NMDA receptor, which might contribute to ischemic tolerance in hippocampal CA1 region of rats.     

The Roles of p38 MAPK/MSK1 Signaling Pathway in the Neuroprotection of Hypoxic Postconditioning Against Transient Global Cerebral Ischemia in Adult Rats

Postconditioning has regenerated interest as a mechanical intervention against cerebral ischemia/reperfusion injury, but its molecular mechanisms remain unknown. We previously reported that hypoxic postconditioning (HPC) ameliorated neuronal death induced by transient global cerebral ischemia (tGCI) in hippocampal CA1 subregion of adult rats. This study tested the hypothesis that p38-mitogen-activated protein kinase (p38 MAPK)/mitogen- and stress-response kinase 1 (MSK1) signaling pathway plays a role in the HPC-induced neuroprotection. Male Wistar rats were subjected to 10 min ischemia induced by applying the four-vessel occlusion method. HPC with 120 min was applied at 24 h after reperfusion. Immunohistochemistry and Western blot were used to detect the expression of phosphorylation of p38 MAPK and MSK1, as well as cleaved caspase-3. We found that HPC induced a significant increase of phosphorylated p38 MAPK and MSK1 in neurons of hippocampal CA1 region and a significant decrease in glial cells after tGCI as well. Furthermore, HPC attenuated caspase-3 cleavation triggered by tGCI in CA1 region. Moreover, p38 MAPK inhibition by SB203580 significantly decreased the phosphorylation of MSK1, increased cleaved caspase-3 expression, and abolished the neuroprotection of HPC. These findings suggested that p38 MAPK/MSK1 signaling axis contributed to HPC-mediated neuroprotection against tGCI, at least in part, by regulating the activation of caspase-3.