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1、Hypothermiainducedneuroprotectionanditsabilitytoattenuate低温诱导的神经保护作用及其能力减弱,1,Hypothermia-induced neuroprotection and its ability to attenuate the down-regulation of GluR2 and GluR1,By Alina SeguraBIO 475Dr. Peter LinSpring Semester,2,Hypothermiainducedneuroprotectionanditsabilitytoattenuate低温诱导的神经保护
2、作用及其能力减弱,Contents,Overview of the CNSGlutamate ReceptorAMPA Receptor HypothermiaSurvival of hippocampal CA1 neuronsDown-regulation of AMPA receptor GluR2 subunitFuture ResearchConclusion,3,Hypothermiainducedneuroprotectionanditsabilitytoattenuate低温诱导的神经保护作用及其能力减弱,Overview of Central Nervous System (
3、CNS),Brain + Spinal CordBrain consists of: forebrain, midbrain, hindbrainForebrain:Telencephalon is its surrounding structurePlays major roles in the sensory perception, learning, memory, and conscience behaviorConsists of the Limbic System,4,Hypothermiainducedneuroprotectionanditsabilitytoattenuate
4、低温诱导的神经保护作用及其能力减弱,The Limbic System,Responsible for the psychological drives, instincts, and emotions,5,Hypothermiainducedneuroprotectionanditsabilitytoattenuate低温诱导的神经保护作用及其能力减弱,The Hippocampus,Transfer of short-term memory to long-term memoryDefined structure Visible layer of pyramidal cellsCA1CA3
5、Dentate Gyrus,6,Hypothermiainducedneuroprotectionanditsabilitytoattenuate低温诱导的神经保护作用及其能力减弱,The Hippocampus,7,Hypothermiainducedneuroprotectionanditsabilitytoattenuate低温诱导的神经保护作用及其能力减弱,Glutamate Receptor,Main component of proteins Present in many tissuesProduced in the bodyPlays essential role in hum
6、an metabolismMajor excitatory neurotransmitter in CNSReceptor plays a vital role in the mediation of excitatory synaptic transmission,8,Hypothermiainducedneuroprotectionanditsabilitytoattenuate低温诱导的神经保护作用及其能力减弱,Glutamate Receptor,Consists 4 or 5 subunits that are subdivided into these receptors:NMDA
7、AMPAKainateAll ionotropic glutamate receptors share common basic structure,9,Hypothermiainducedneuroprotectionanditsabilitytoattenuate低温诱导的神经保护作用及其能力减弱,Ionotropic Glutamate Receptors,10,Hypothermiainducedneuroprotectionanditsabilitytoattenuate低温诱导的神经保护作用及其能力减弱,Synaptic Transmission,Synthesis of glut
8、amate (GLU) from glutamineTransport and storageRelease of GLU by exocytosisBinding of GLU to antagonist receptorsBinding to presynaptic receptorsReuptakeDegradation,11,Hypothermiainducedneuroprotectionanditsabilitytoattenuate低温诱导的神经保护作用及其能力减弱,AMPA Receptor,Alpha-amino-3-hydroxy-5-methyl-4-isoxazolep
9、ropionic acidComposed of GluR2-4Channels are impermeable to calcium ionsHowever, the calcium permeability of the GluR2 subunit is determined by the post-transcriptional editing of the GluR2 mRNA,12,Hypothermiainducedneuroprotectionanditsabilitytoattenuate低温诱导的神经保护作用及其能力减弱,AMPA Receptor,Change in sin
10、gle amino acid in the TM2 region from glutamine (Q) to arginine (R)GluR2 (Q) is calcium permeable,13,Hypothermiainducedneuroprotectionanditsabilitytoattenuate低温诱导的神经保护作用及其能力减弱,Ischemia,Condition in which tissues and organs suffer inadequate blood flow in particular or widespread area of the CNSGloba
11、l ischemia models are used to create a reproducible region of ischemia in the rodent or gerbil forebrain In gerbils, the methods used for these experiments are bilateral carotid occlusion with hypotension or ligation of both vertebral arteries and transient occlusion of both carotids, with or withou
12、t hypotension,14,Hypothermiainducedneuroprotectionanditsabilitytoattenuate低温诱导的神经保护作用及其能力减弱,Hypothermia,When body temperature reaches temperatures lower than 35CMild (32.2 to 35C)Moderate (28 to 32C)Severe (below 28C)Induces some neuroprotective actions,15,Hypothermiainducedneuroprotectionanditsabil
13、itytoattenuate低温诱导的神经保护作用及其能力减弱,Hypothermia: Neuroprotective mechanisms,Hypothermiainducedneuroprotectionanditsabilitytoattenuate低温诱导的神经保护作用及其能力减弱,16,Colbourne, F., S.Y. Grooms, R.S. Zukin, A.M. Buschan, M.L.Bennett. 2003. Hypothermia rescues hippocampal CA1 neurons and attenuates down-regulation of
14、 the AMPA receptor GluR2 subunit after forebrain ischemia. Proceedings of the National Academy of Science in the United of America. 100: 2906-2910.,Hypothermia rescues hippocampal CA1 neurons and attenuates down-regulation of the AMPA receptor GluR2 subunit after forebrain ischemia,17,Hypothermiaind
15、ucedneuroprotectionanditsabilitytoattenuate低温诱导的神经保护作用及其能力减弱,Purpose,Colbourne et al. (2003) wanted to examine the mechanisms underlying hypothermia-induced neuroprotection.,18,Hypothermiainducedneuroprotectionanditsabilitytoattenuate低温诱导的神经保护作用及其能力减弱,Brain Temperature,All gerbils were implanted a t
16、elemetry probe that measured temperature of dorsal striatumGlobal ischemia in the gerbil was produced 2 days after probe insertion by the occlusion of the common carotid arteries.Temperature was maintained near normothermia (36.0C),19,Hypothermiainducedneuroprotectionanditsabilitytoattenuate低温诱导的神经保
17、护作用及其能力减弱,Brain Temperature,In order to observe the preservation of the CA1 neurons, they measured the postischemic temperature.Four groups were examined:Sham Sham + HypothermiaIschemiaIschemia + Hypothermia,20,Hypothermiainducedneuroprotectionanditsabilitytoattenuate低温诱导的神经保护作用及其能力减弱,Figure 1: Brai
18、n Temperature,21,Hypothermiainducedneuroprotectionanditsabilitytoattenuate低温诱导的神经保护作用及其能力减弱,Results,22,Hypothermiainducedneuroprotectionanditsabilitytoattenuate低温诱导的神经保护作用及其能力减弱,Results,Figure 1C, gerbils subjected to ischemia without treatment were allowed to survive for 7 days to illustrate the lo
19、ss of GluR1 and GluR2 mRNA in CA1 neurons that experienced cell death.Prolonged, delayed hypothermia induced 1 hour after ischemia and maintained for 48 hours rescued most CA1 pyramidal neurons as shown in Figure 1D.,23,Hypothermiainducedneuroprotectionanditsabilitytoattenuate低温诱导的神经保护作用及其能力减弱,How d
20、oes this happen?,In order to examine the mechanisms by which prolonged postischemic cooling rescues neurons, Colbourne et al. looked at the relative abundance of GluR2 and GluR1 mRNA .This was assessed by a technique called in situ hybridization,24,Hypothermiainducedneuroprotectionanditsabilitytoatt
21、enuate低温诱导的神经保护作用及其能力减弱,In Situ Hybridization,method of localizing and detecting specific mRNA sequences in preserved tissues sections or cell preparations by hybridizing the complimentary strand of a nucleotide probe to the sequence of GluR2 mRNA expression.,25,Hypothermiainducedneuroprotectionandi
22、tsabilitytoattenuate低温诱导的神经保护作用及其能力减弱,Methods of In Situ Hybridization,Gerbils were decapitatedFrozen cryosections were obtained prior to hybridizationOligonucleotide probe (RNA probe) was inserted35S-labeled GluR1GluR2 riboprobesCryosections were hybridized,26,Hypothermiainducedneuroprotectionandit
23、sabilitytoattenuate低温诱导的神经保护作用及其能力减弱,Methods continued,To quantify mRNA expression of GluR1 and GluR2 film autoradiograms were performed.Sham (2 day survival)Sham + Hypothermia (2 and 7 day survival times)Global Ischemia (2 and 7 day survival times)Global Ischemia + Hypothermia (2 and 7 day survival
24、 times)They are reported as a ratio of optical densities and were averaged for 2 to 4 sections per gerbil for 4 to 6 gerbils.Optical densities values were expressed in SD,27,Hypothermiainducedneuroprotectionanditsabilitytoattenuate低温诱导的神经保护作用及其能力减弱,28,Hypothermiainducedneuroprotectionanditsabilityto
25、attenuate低温诱导的神经保护作用及其能力减弱,Results,Figure 2A and 2B showed that untreated forebrain ischemia induced a pronounced decrease in GluR2 mRNA expression in the CA1 sector at 2 daysAlso, showed near total loss at 7 days The decrease in GluR2 at 2 days in ischemic gerbils treated with delayed cooling was s
26、ignificantly attenuated,29,Hypothermiainducedneuroprotectionanditsabilitytoattenuate低温诱导的神经保护作用及其能力减弱,Results continued,In GluR1 mRNA expression in CA1 at 2 days, forebrain ischemia showed a slight, but significant decreaseAt 7 days, there was complete loss of CA1 neuronsThere is a small, but signif
27、icant down-regulation of GluR1 mRNA expression,30,Hypothermiainducedneuroprotectionanditsabilitytoattenuate低温诱导的神经保护作用及其能力减弱,Expression of GluR2 and GluR1,Emulsion-dipped (Cresyl violet-stained) coronal sections of the hippocampal CA1 zone were obtained to also show the expression of GluR1 and GluR2
28、A-D demonstrate GluR2E-H demonstrate GluR1Arrows show silver grains that overly individual pyramidal cells in CA1,31,Hypothermiainducedneuroprotectionanditsabilitytoattenuate低温诱导的神经保护作用及其能力减弱,32,Hypothermiainducedneuroprotectionanditsabilitytoattenuate低温诱导的神经保护作用及其能力减弱,Results,In slide B for GluR2 t
29、he number of silver grains is reduced after ischemia suggesting decrease in GluR2 mRNA expresssion Reduction was also seen in slide C for GluR2 (ischemia+hypothermia) at 2 daysComplete recovery was marked at 7 daysHowever, in GluR1 mRNA expression in CA1 at 2 days after untreated ischemia (slide F)w
30、as prevented by hypothermia,33,Hypothermiainducedneuroprotectionanditsabilitytoattenuate低温诱导的神经保护作用及其能力减弱,Future Studies,There are still many questions underlying the mechanisms by which hypothermia causes a variety of responses in ischemic brain that might confer neuroprotection.Therefore, future s
31、tudies delineating these mechanisms are necessary.,34,Hypothermiainducedneuroprotectionanditsabilitytoattenuate低温诱导的神经保护作用及其能力减弱,Conclusion,Prolonged postischemic hypothermia significantly attenuated the decline in GluR2 mRNA and promoted its full recovery. Changes to GluR1 were completely prevented. Data suggest that delayed and prolonged hypothermia reduces ischemic CA1 damage through reducing the formation of modified GluR2-lacking AMPARs, which otherwise leads to influx of toxic levels of Ca2+ and possibly Zn2+.,