Lessons from nature: signalling cascades associated with vertebrate brain anoxic survival.

NEW FINDINGS: What is the topic of this review? Although the mammalian brain is exquisitely sensitive to hypoxia, some turtles survive complete anoxia by decreasing metabolic demand to match reduced energy supply. These animal models may help to elucidate neuroprotective mechanisms and reveal novel therapeutic targets for diseases of oxygen deprivation. What advances does it highlight? The mitogen-activated protein kinases (MAPKs) are part of the suite of adaptive responses to anoxia that are modulated by adenosine, a 'retaliatory metabolite' released in early anoxia. In anoxic turtle neurons, upregulation of pro-survival Akt and extracellular signal-regulated kinase 1/2 and suppression of the p38MAPK and JNK pathways promote cell survival, as does the anoxic- and post-anoxic upregulation of the antioxidant methionine sulfoxide reductase. Mammalian neurons undergo rapid degeneration when oxygen supply is curtailed. Neuroprotective pathways are induced during hypoxia/ischaemia, but their analysis is complicated by concurrent pathological events. Survival mechanisms can be investigated in anoxia-tolerant freshwater turtle species, which survive oxygen deprivation and post-anoxic reoxygenation by entrance into a state of reversible hypometabolism. Many energy-demanding processes are suppressed, including ion flux and neurotransmitter release, whereas cellular protective mechanisms, including certain mitogen-activated protein kinases (MAPKs), are upregulated. This superfamily of serine/threonine kinases plays a significant role in vital cellular processes, including cell proliferation, differentiation, stress adaptation and apoptosis in response to external stimuli. Here, we report that neuronal survival relies on robust co-ordination between the major signalling cascades, with upregulation of the pro-survival Akt and extracellular signal-regulated kinase 1/2 and suppression of the p38MAPK and JNK pathways. Other protective responses, including the upregulation of heat shock proteins and antioxidants, allow the turtle brain to abrogate potential oxidative stress upon reoxygenation.

Upregulation of Hsp72 mediates anoxia/reoxygenation neuroprotection in the freshwater turtle via modulation of ROS.

The neuroprotective role of Hsp72 has been demonstrated in several ischemic/stroke models to occur primarily through mediation of apoptotic pathways, and a number of heat shock proteins are upregulated in animal models capable of extended anoxic survival. In the present study, we investigated the role of Hsp72 on cell death and apoptotic regulators in one anoxia tolerant model system, the freshwater turtle Trachemys scripta. Since Hsp72 is known to regulate apoptosis through interactions with Bcl-2, we manipulated the levels of Hsp72 and Bcl-2 with siRNA in neuronally enriched primary cell cultures and examined downstream effects. The knockdown of either Hsp72 or Bcl-2 induced cell death during anoxia and reoxygenation. Knockdown of Bcl-2 resulted in increases in apoptotic markers and increased ROS levels 2-fold. However, significant knockdown of Hsp72 did not have any effect on the expression of key mitochondrial apoptotic regulators such as Cytochrome c and caspase-3. Hsp72 knockdown however significantly increased apoptosis inducing factor in both anoxia and reoxygenation and resulted in a six-fold induction of hydrogen peroxide levels. These findings suggest that the neuroprotection offered by Hsp72 in the anoxia/reoxygenation tolerant turtle is through the mediation of ROS levels and not through modulation of caspase-dependent pathways.

The E-box binding factors Max/Mnt, MITF, and USF1 act coordinately with FoxO to regulate expression of proapoptotic and cell cycle control genes by phosphatidylinositol 3-kinase/Akt/glycogen synthase kinase 3 signaling.

Phosphatidylinositol (PI) 3-kinase/Akt signaling plays a critical role in cell proliferation and survival, partly by regulation of FoxO transcription factors. Previous work using global expression profiling indicated that inhibition of PI 3-kinase in proliferating cells led to induction of genes that promote cell cycle arrest and apoptosis. The upstream regulatory regions of these genes had binding sites not only for FoxO, but also for Myc/Max transcription factors. In the present study, we have addressed the role of Myc family members and related E-box-binding proteins in the regulation of these genes. Chromatin immunoprecipitations and RNA interference indicated that transcription was repressed by Max-Mnt-Sin3a-histone deacetylase complexes in proliferating cells. Inhibition of PI 3-kinase led to a loss of Max/Mnt binding and transcriptional induction by MITF and USF1, as well as FoxO. Both MITF and USF1 were activated by glycogen synthase kinase (GSK) 3, with GSK3 phosphorylation sites on USF1 identified as the previously described activating site threonine 153 as well as serine 186. siRNA against MITF as well as against FoxO3a protected cells from apoptosis following PI 3-kinase inhibition. These results define a novel E-box-regulated network that functions coordinately with FoxO to regulate transcription of apoptotic and cell cycle regulatory genes downstream of PI 3-kinase/Akt/GSK3 signaling.

Neuroprotective signaling pathways are modulated by adenosine in the anoxia tolerant turtle.

Cumulative evidence shows a protective role for adenosine A1 receptors (A1R) in hypoxia/ischemia; A1R stimulation reduces neuronal damage, whereas blockade exacerbates damage. The signal transduction pathways may involve the mitogen-activated protein kinase (MAPK) pathways and serine/threonine kinase (AKT), with cell survival depending on the timing and degree of upregulation of these cascades as well as the balance between pro-survival and pro-death pathways. Here, we show in vitro that extracellular signal-regulated kinase (ERK1/2) and phosphatidylinositol 3-kinase (PI3-K/AKT) activation is dependent on A1R stimulation, with further downstream effects that promote neuronal survival. Phosphorylated ERK1/2 (p-ERK) and AKT (p-AKT) as well as Bcl-2 are upregulated in anoxic neuronally enriched primary cultures from turtle brain. This native upregulation is further increased by the selective A1R agonist 2-chloro-N-cyclopentyladenosine (CCPA), whereas the selective antagonist 8-cyclopentyl-1,3-dihydropylxanthine (DPCPX) decreases p-ERK and p-AKT expression. Conversely, A1R antagonism resulted in increases in phosphorylated JNK (p-JNK), p38 (p-p38), and Bax. As pathological and adaptive changes occur simultaneously during anoxia/ischemia in mammalian neurons, the turtle provides an alternative model to analyze protective mechanisms in the absence of evident pathologies.

Role of neuroglobin in regulating reactive oxygen species in the brain of the anoxia-tolerant turtle Trachemys scripta.

Neuroglobin (Ngb) is an oxygen binding heme protein found in nervous tissue with a yet unclear physiological and protective role in the hypoxia-sensitive mammalian brain. Here we utilized in vivo and in vitro studies to examine the role of Ngb in anoxic and post-anoxic neuronal survival in the freshwater turtle. We employed semiquantitative RT-PCR and western blotting to analyze Ngb mRNA and protein levels in turtle brain and neuronally enriched cultures. Ngb expression is strongly up-regulated by hypoxia and post-anoxia reoxygenation but increases only modestly in anoxia. The potential neuroprotective role of Ngb in this species was analyzed by knocking down Ngb using specific small interfering RNA. Ngb knockdown in neuronally enriched cell cultures resulted in significant increases in H(2)O(2) release compared to controls but no change in cell death. Cell survival may be linked to activation of other protective responses such as the extracellular regulated kinase transduction pathway, as phosphorylated extracellular regulated kinase levels in anoxia were significantly higher in Ngb knockdown cultures compared to controls. The greater expression of Ngb when reactive oxygen species are likely to be high, and the increased susceptibility of neurons to H(2)O(2) release and external oxidative stress in knockdown cultures, suggests a role for Ngb in reducing reactive oxygen species production or in detoxification, though it does not appear to be of primary importance in the anoxia tolerant turtle in the presence of compensatory survival mechanisms.

Brain response correlates of decisional capacity in schizophrenia: a preliminary FMRI study.

The capacity of schizophrenia patients to make decisions regarding research consent relates to neurocognition, but the exact nature of the relationship is unclear. The authors examined the correlation of scores on the MacArthur Competence Assessment Tool for Clinical Research with functional magnetic resonance imaging brain response during a verbal learning task. Understanding of a consent form correlated with activation of the right hippocampus during verbal learning and with brain response in a large area that included the bilateral parahippocampus, cerebellum, and thalamus. Reasoning scores were not significantly related to brain activation. Understanding deficits during informed consent relates to particular brain abnormalities among schizophrenia patients.

Suppression of reactive oxygen species production enhances neuronal survival in vitro and in vivo in the anoxia-tolerant turtle Trachemys scripta.

Hypoxia-ischemia with reperfusion is known to cause reactive oxygen species-related damage in mammalian systems, yet, the anoxia tolerant freshwater turtle is able to survive repeated bouts of anoxia/reoxygenation without apparent damage. Although the physiology of anoxia tolerance has been much studied, the adaptations that permit survival of reoxygenation stress have been largely ignored. In this study, we examine ROS production in the turtle striatum and in primary neuronal cultures, and examine the effects of adenosine (AD) on cell survival and ROS. Hydroxyl radical formation was measured by the conversion of salicylate to 2,3-dihydroxybenzoic acid (2,3-DHBA) using microdialysis; reoxygenation after 1 or 4 h anoxia did not result in increased ROS production compared with basal normoxic levels, nor did H(2)O(2) increase after anoxia/reoxygenation in neuronally enriched cell cultures. Blockade of AD receptors increased both ROS production and cell death in vitro, while AD agonists decreased cell death and ROS. As turtle neurons proved surprisingly susceptible to externally imposed ROS stress (H(2)O(2)), we propose that the suppression of ROS formation, coupled to high antioxidant levels, is necessary for reoxygenation survival. As an evolutionarily selected adaptation, the ability to suppress ROS formation could prove an interesting path to investigate new therapeutic targets in mammals.

Gene transcription of neuroglobin is upregulated by hypoxia and anoxia in the brain of the anoxia-tolerant turtle Trachemys scripta.

Neuroglobin is a heme protein expressed in the vertebrate brain in mammals, fishes, and birds. The physiological role of neuroglobin is not completely understood but possibilities include serving as an intracellular oxygen-carrier or oxygen-sensor, as a terminal oxidase to regenerate NAD(+) under anaerobic conditions, or involvement in NO or ROS metabolism. As the vertebrate nervous system is particularly sensitive to hypoxia, an intracellular protein that helps sustain cellular respiration would aid hypoxic survival. However, the regulation of Neuroglobin (Ngb) under conditions of varying oxygen is controversial. This study examines the regulation of Ngb in an anoxia-tolerant vertebrate under conditions of hypoxia and anoxia. The freshwater turtle Trachemys scripta can withstand complete anoxia for days, and adaptations that permit neuronal survival have been extensively examined. Turtle neuroglobin specific primers were employed in RT-PCR for determining the regulation of neuroglobin mRNA expression in turtles placed in normoxia, hypoxia (4 h), anoxia (1 and 4 h), and anoxia-reoxygenation. Whole brain expression of neuroglobin is strongly upregulated by hypoxia and post-anoxic-reoxygenation in T. scripta, with a lesser degree of upregulation at 1 and 4 h anoxia. Our data implicate neuroglobin in mediating brain anoxic survival.

Atypical antipsychotics in elderly patients with dementia or schizophrenia: review of recent literature.

Atypical antipsychotics have become a common pharmacologic option for the treatment of various psychiatric and behavioral symptoms in older adults, although these medications have been officially approved by the U.S. Food and Drug Administration for use only in schizophrenia and bipolar disorder. Despite the widespread use of these agents, there is a relative shortage of rigorously conducted trials. This review focuses on recently published randomized, blinded, controlled trials involving the use of atypical antipsychotics in elderly patients with dementia (n = 9) or schizophrenia (n = 3), with some discussion of published large, open-label studies and a few unpublished controlled trials. In general, the studies of patients with dementia reported modest efficacy of atypical antipsychotics when compared to placebo and conventional antipsychotics. In addition, an advantage in terms of motor side effects was consistently noted with atypical antipsychotics when compared to conventional antipsychotics. The studies have also shown, however, a greater risk of mortality and adverse cerebrovascular events with several of these agents than with placebo in individuals with dementia. There are insufficient data comparing atypical antipsychotics to one another. In the trials involving elderly persons with schizophrenia, atypical antipsychotics were associated with significant improvements in psychopathology; differences in efficacy among atypical antipsychotics were unclear. A careful consideration of the risk-benefit ratio of atypical antipsychotics, as well as that of available alternative treatments, is needed for each individual elderly patient. Clinical judgment, caution, and consent should be the watchwords in this area of psychopharmacology.

Generalized cognitive impairments, ability to perform everyday tasks, and level of independence in community living situations of older patients with psychosis.

OBJECTIVE: This study examined the relationship between performance of instrumental activities of daily living, as measured with the University of California, San Diego, Performance-Based Skills Assessment (UPSA), and measures of cognitive functioning and independence in the community living situation of older outpatients with psychotic disorders. METHOD: One hundred eleven middle-aged and elderly outpatients with primary psychotic disorders were administered the UPSA, the Mattis Dementia Rating Scale, and standardized measures of psychopathology. Independence in the community living situation was rated on a scale from 1 to 5, with 5 representing the highest level of independence, such as living alone in an apartment or house. Ability in seven domains of neuropsychological functioning was assessed in 67 participants. RESULTS: Total scores on the University of California, San Diego, Performance-Based Skills Assessment were significantly correlated with total and subscale scores on the Mattis Dementia Rating Scale, level of independence in the community living situation, and scores on the neuropsychological measures in the subset of patients who underwent neuropsychological testing. Lower scores on the Mattis Dementia Rating Scale memory subscale and more severe negative symptoms were significantly associated with worse performance on the skills assessment. Performance on the skills assessment contributed uniquely to prediction of independence in the community living situation. CONCLUSIONS: Generalized cognitive abilities are associated with everyday functioning capacity in older patients with psychosis. Reduction of cognitive deficits and negative symptoms may improve patients' ability to function independently in the community.

Treatment-related decision-making capacity in middle-aged and older patients with psychosis: a preliminary study using the MacCAT-T and HCAT.

The authors conducted a preliminary examination of treatment-related decision-making capacity (DMC) in middle-aged and older psychosis patients. The MacArthur Competency Assessment Tool for Treatment (MacCAT-T) was administered to measure DMC among 16 patients with psychotic disorders and without dementia (mean age: 54.6 [SD 7.2] years). The impact of repeated learning trials on patients' understanding was assessed with the Hopkins Competency Assessment Test (HCAT). Subjects were also assessed with the Positive And Negative Syndrome Scale and Mattis' Dementia Rating Scale. There was a wide range of performance on the MacCAT-T; the levels and range of MacCAT-T scores were similar to those previously reported among younger acutely hospitalized patients in the MacCAT-T validation study. Patients' understanding of disclosed material on the HCAT improved significantly over repeated presentations. Although there were no statistically significant correlations observed between DMC and demographic characteristics, severity of symptoms, or global cognitive deficits, the sample size provided limited power to detect such associations. Results point to the importance of assessing DMC and to the potential modifiability of initial difficulties in understanding.

The California Scale of Appreciation: a new instrument to measure the appreciation component of capacity to consent to research.

Capacity to consent is one of the linchpins of the ethical conduct of clinical care and research, and it needs to be reliably measured. The authors describe the development of a new measure of the "appreciation" component of capacity, the California Scale of Appreciation (CSA), 18 items rated according to the concept of "patently false belief" (a belief that is grossly improbable); 39 patients with schizophrenia or a related psychotic disorder (27 outpatients and 12 inpatients) and 15 normal-comparison subjects participated. Each subject's audiotaped interview was rated by three evaluators. Answers to each item were scored as "capable," "incapable," or "uncertain capacity." Also, each subject was given an overall rating of one of these three categories by each rater. Total scores on the CSA were calculated and correlated with scores on standardized instruments for assessing psychopathology and cognitive impairment. The mean total CSA score was significantly lower in the patients than in the normal-comparison subjects; however, a majority of the patients were found to be fully "capable" on the CSA. The CSA is a potentially useful instrument for measuring the appreciation component of capacity in persons with psychotic disorders. Its generalizability to other patient populations and to other types of protocols needs to be determined.