Biomedical subjects

Leslie Thomas Buck

Publications and source records attributed to Leslie Thomas Buck.

5 recordsLinked to original sources

PubMed2005

Calcium and protein phosphatase 1/2A attenuate N-methyl-D-aspartate receptor activity in the anoxic turtle cortex.

Excitotoxic cell death (ECD) is characteristic of mammalian brain following min of anoxia, but is not observed in the western painted turtle following days to months without oxygen. A key event in ECD is a massive increase in intracellular Ca(2+) by over-stimulation of N-methyl-d-aspartate receptors (NMDARs). The turtle's anoxia tolerance may involve the prevention of ECD by attenuating NMDAR-induced Ca(2+) influx. The goal of this study was to determine if protein phosphatases (PPs) and intracellular calcium mediate reductions in turtle cortical neuron whole-cell NMDAR currents during anoxia, thereby preventing ECD. Whole-cell NMDAR currents did not change during 80 min of normoxia, but decreased 56% during 40 min of anoxia. Okadaic acid and calyculin A, inhibitors of serine/threonine PP1 and PP2A, potentiated NMDAR currents during normoxia and prevented anoxia-mediated attenuation of NMDAR currents. Decreases in NMDAR activity during anoxia were also abolished by inclusion of the Ca(2+) chelator -- BAPTA and the calmodulin inhibitor -- calmidazolium. However, cypermethrin, an inhibitor of the Ca(2+)/calmodulin-dependent PP2B (calcineurin), abolished the anoxic decrease in NMDAR activity at 20, but not 40 min suggesting that this phosphatase might play an early role in attenuating NMDAR activity during anoxia. Our results show that PPs, Ca(2+) and calmodulin play an important role in decreasing NMDAR activity during anoxia in the turtle cortex. We offer a novel mechanism describing this attenuation in which PP1 and 2A dephosphorylate the NMDAR (NR1 subunit) followed by calmodulin binding, a subsequent dissociation of alpha-actinin-2 from the NR1 subunit, and a decrease in NMDAR activity.

Animals↗

PubMed2004

Adenosine as a signal for ion channel arrest in anoxia-tolerant organisms.

Certain freshwater turtles and fish are extremely anoxia-tolerant, capable of surviving hours of anoxia at high temperatures and weeks to months at low temperatures. There is great interest in understanding the cellular mechanisms underlying anoxia-tolerance in these groups because they are anoxia-tolerant vertebrates and because of the far-reaching medical benefits that would be gained. It has become clear that a pre-condition of prolonged anoxic survival must involve the matching of ATP production with ATP utilization to maintain stable ATP levels during anoxia. In most vertebrates, anoxia leads to a severe decrease in ATP production without a concomitant reduction in utilization, which inevitably leads to the catastrophic events associated with cell death or necrosis. Anoxia-tolerant organisms do not increase ATP production when faced with anoxia, but rather decrease utilization to a level that can be met by anaerobic glycolysis alone. Protein synthesis and ion movement across the plasma membrane are the two main targets of regulatory processes that reduce ATP utilization and promote anoxic survival. However, the oxygen sensing and biochemical signaling mechanisms that achieve a coordinated reduction in ATP production and utilization remain unclear. One candidate-signaling compound whose extracellular concentration increases in concert with decreasing oxygen availability is adenosine. Adenosine is known to have profound effects on various aspects of tissue metabolism, including protein synthesis, ion pumping and permeability of ion channels. In this review, I will investigate the role of adenosine in the naturally anoxia-tolerant freshwater turtle and goldfish and give an overview of pathways by which adenosine concentrations are regulated.

Adaptation, Physiological↗

PubMed2003

Gap junctions do not underlie changes in whole-cell conductance in anoxic turtle brain.

An acute reduction in cell membrane permeability could provide an effective strategy to prolong anoxic survival. A previous study has shown that in the western painted turtle whole-cell neuronal conductance (G(w)) decreases during anoxia, which may be mediated by the activation of adenosine A(1) receptors and calcium. Reduction in G(w) is thought to be the result of ion channel closure, but closure of gap junctions could also be responsible for this phenomenon. In our study, antibody staining of connexin 32 and 43 (Cx32 and Cx43) suggested the presence of gap junctional components in the turtle cortex. To examine if gap junctions were involved in the previously measured anoxic decrease in G(w), neuronal connectivity was assessed through the measurement of whole-cell capacitance (C(w)). Turtle cortical sheets were perfused with normoxic (95%O(2)/5%CO(2)), anoxic (95%N(2)/5%CO(2)), high calcium (4 mM) and adenosine (200 microm) artificial cerebral spinal fluid (aCSF). No significant change in C(w) was observed under any of the above conditions. However, during hypo-osmotic aCSF perfusion C(w) decreased significantly, with the lowest value of 50+/-10.4 pF (P<0.05) occurring at 30 min. To visualize changes in gap junction permeability lucifer yellow was loaded into turtle neurons during normoxic, anoxic, 0 calcium, hypo-osmotic, cold shock, (+)-isoproterenol, nitric oxide donor S-nitoso-acetyl penicillamine, and 8-bromo-guanosine 3',5'-cyclic monophosphate aCSF perfusion. Dye propagation was only observed in 3 of 20 cold shock experiments (4 degrees C). We conclude that gap junctions are not involved in the acute reduction in G(w) previously observed during anoxia and that our results support the hypothesis that ion channel arrest is involved.

Animals↗

PubMed2003

Tissue-specific expression of inducible and constitutive Hsp70 isoforms in the western painted turtle.

Expression of Hsp73 and Hsp72 in four tissues of the naturally anoxia-tolerant western painted turtle (Chrysemys picta) was investigated in response to a 24 h forced dive and following 1 h recovery. Of the tissues examined, brain and liver displayed approximately threefold and sevenfold higher basal Hsp73 expression than heart and skeletal muscle. Basal Hsp72 expression was relatively low in all tissues examined. After the 24 h forced dive and 1 h recovery, Hsp73 expression did not differ significantly from basal expression with the exception of liver, where expression decreased significantly after 1 h recovery. Hsp72 expression was unchanged in liver following a 24 h dive; however, it increased twofold in brain and threefold in heart and skeletal muscle. Dive-induced Hsp72 expression was found to correlate inversely with basal Hsp73 expression. Following 1 h recovery, Hsp72 expression was significantly elevated in all tissues above levels in dived animals. These data indicate a tissue-specific pattern of Hsp73 and Hsp72 expression in the western painted turtle during both unstressed and stressed conditions.

Animals↗

PubMedDate not supplied

Effect of anoxia and pharmacological anoxia on whole-cell NMDA receptor currents in cortical neurons from the western painted turtle.

The mammalian brain undergoes rapid cell death during anoxia that is characterized by uncontrolled Ca(2+) entry via N-methyl-D-aspartate receptors (NMDARs). In contrast, the western painted turtle is extremely anoxia tolerant and maintains close-to-normal [Ca(2+)](i) during periods of anoxia lasting from days to months. A plausible mechanism of anoxic survival in turtle neurons is the regulation of NMDARs to prevent excitotoxic Ca(2+) injury. However, studies using metabolic inhibitors such as cyanide (NaCN) as a convenient method to induce anoxia may not represent a true anoxic stress. This study was undertaken to determine whether turtle cortical neuron whole-cell NMDAR currents respond similarly to true anoxia with N(2) and to NaCN-induced anoxia. Whole-cell NMDAR currents were measured during a control N(2)-induced anoxic transition and a control NaCN-induced transition. During anoxia with N(2) normalized, NMDAR currents decreased to 35.3%+/-10.8% of control values. Two different NMDAR current responses were observed during NaCN-induced anoxia: one resulted in a 172%+/-51% increase in NMDAR currents, and the other was a decrease to 48%+/-14% of control. When responses were correlated to the two major neuronal subtypes under study, we found that stellate neurons responded to NaCN treatment with a decrease in NMDAR current, while pyramidal neurons exhibited both increases and decreases. Our results show that whole-cell NMDAR currents respond differently to NaCN-induced anoxia than to the more physiologically relevant anoxia with N(2).

Animals↗