Exocytosis And Iii. Oxidative Stress
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Exocytosis And Iii. Oxidative Stress

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Exocytosis And Iii. Oxidative Stress Exocytosis And Iii. Oxidative Stress Presentation Transcript

  • Neurons and Oxidative stress in Brain: “Claiming for the Right Supply of Blood” Christian Amatore Ecole Normale Supérieure, Département de Chimie UMR CNRS-ENS-UPMC 8640 "PASTEUR" Paris - France
  • Adapted from: http://www.abcam/neuroscience/
    • Imaging Neurotransmission in the Brain by IRMf:
    Ecoute subjective ou analytique d’une même pièce de musique par le même sujet.  Dept. Neurology and Radiology, Münster. IRMf d’un sujet adulte de 24 ans durant une tâche de génération de mots .  Dept. Neurology and Radiology, Münster. IRMf. Photo: Jeff Miller.  University of Wisconsin-Madison. Subjective Analytical
  • d d +  d
    • Modulation of Complex Biological Assemblies Interrelations
    • Control of Blood Supply in Brain by Local Release of NO :
    • Control of Blood Supply in Brain by Local Release of NO ?
    Rat Brain Slice 300µm. Collaboration with J Rossier, A Rancillac & B Cauli, UMR CNRS 7637, ESPCI, Paris. Unpublished results. 2004.
    • Oxidative Stress and Neurotransmission:
    • Control of Blood Supply by Dilatation of Blood Vessels :
    muscle cells endothelial cells tissue lumen
    • Control of Blood Supply by Dilatation of Blood Vessels :
    lumen guanylate cyclases phosphokinases proteins phosphorylation
    • Oxidative Stress and Neurotransmission:
    NO°
    • NO Release in Aerobic Cells
    • NO Synthases:
    N H 2 2 + N H 2 N H N N H 2 O N H H 3 N + O O - N H H 3 N + O O - N H H 3 N + O O - H O O 2 + e O 2 N O 2e L-Arginine N-hydroxy-arginine Citrulline
    • Modulation of Complex Biological Assemblies Interrelations
    • Control of Blood Supply in Brain by Local Release of NO :
    • Control of Blood Supply in Brain by Local Release of NO :
    Rat Brain sagittal cut (300µm; vibratome) stellate neuron cerebellum
    • Modulation of Complex Biological Assemblies Interrelations
    20 µm
    • Modulation of Complex Biological Assemblies Interrelations
    • Molecular Characterization of NOS in Stellate Neuron
    Rat Brain Slice 300µm. Collaboration with J Rossier, A Rancillac & B Cauli, UMR CNRS 7637, ESPCI, Paris. submitted Patch Clamp onto Neuron Body: stellate neuron body patch-clamp pipette blood capillary vessel
    • Molecular Characterization of NOS in Stellate Neuron
    Rat Brain Slice 300µm. Collaboration with J Rossier, A Rancillac & B Cauli, UMR CNRS 7637, ESPCI, Paris. submitted Characterization of NOS (RT PCR) Patch Clamp onto Neuron Body: stellate neuron body patch-clamp pipette blood capillary vessel
    • Platinized Carbon Fiber Ultramicroelectrodes
    10-12 µm 1-5 µm glass cases insulating polymer platinized surfaces 5 µm 0 0.2 0.4 0.6 0.8 1 1.2 200 400 600 800 Current (a.u.) E / mV vs. SSCE NO° / 1µM
    • Platinized Carbon Fiber Ultramicroelectrodes
    • Modulation of Complex Biological Assemblies Interrelations
    • Control of Blood Supply in Brain by Local Release of NO :
    • Control of Blood Supply in Brain by Local Release of NO :
    Rat Brain Slice 300µm. Collaboration with J Rossier, A Rancillac & B Cauli, UMR CNRS 7637, ESPCI, Paris. Unpublished results. 2004. artificial brain medium based on Ringer buffer optical microscope peristaltic pump patch-clamp rat brain slice O 2 & CO 2 potentiostat potential CAMERA camera white and IR light source for differential contrast NO-ultramicroelectrode
    • Modulation of Complex Biological Assemblies Interrelations
    • Test 1: Vessel Stimulation by External Local Chemical Release of NO°
    5 µm 20 pA 5 µm (  5% ) 5 min Rat Brain Slice 300µm. 0.65 V vs. SSCE. Collaboration with J Rossier, A Rancillac & B Cauli, UMR CNRS 7637, ESPCI, Paris. Unpublished results. 2004. time vessel diameter current DEANOOate 100 µm 0.5 µM NO° 0.62 V vs. Ag/AgCl diazeniumdiolate - d
  • Rat Brain Slice 300µm. 0.65 V vs. SSCE. Collaboration with J Rossier, A Rancillac & B Cauli, UMR CNRS 7637, ESPCI, Paris. Unpublished results. 2004.
    • Modulation of Complex Biological Assemblies Interrelations
    • Test 2: Stimulation via Chemical Stimulation of Stellate Neuron by NMDA
    0.62 V vs. Ag/AgCl NMDA NO° Star neurons NOsynthase d current NMDA 100 µM 5 pA 0.15 µM NO°
    • Controls:
    • TTX (  ) : tetrodotoxin (blocking of ion channels)
    • LNAME (  ) : Nitro L Arginine Methyl Ester
    vessel diameter time 2 µm (  2% ) 5 min
    • Electrophysiological Stimulation with Patch-Clamp Pipette
    • Modulation of Complex Biological Assemblies Interrelations
    NO° Release Rat Brain Slice 300µm. 0.65 V vs. SSCE. Collaboration with J Rossier, A Rancillac & B Cauli, UMR CNRS 7637, ESPCI, Paris. Unpublished results. 2004. ~ V Action Potential Firing L-NAME (1 mM) TTX (1 µM) 0 5 10 -2 0 8 10 6 4 2 Patch-Clamp Stim -0.04 0.04 0.08 0.12 0.16 0 Current (pA) NO° flux (fmol/s)
    • Electrophysiological Stimulation with Patch-Clamp Pipette
    • Modulation of Complex Biological Assemblies Interrelations
    Rat Brain Slice 300µm. 0.65 V vs. SSCE. Collaboration with J Rossier, A Rancillac & B Cauli, UMR CNRS 7637, ESPCI, Paris. Unpublished results. 2004.
    • Electrophysiological Stimulation with Patch-Clamp Pipette
    • Modulation of Complex Biological Assemblies Interrelations
    NO° Release and Vessel Dilatation Rat Brain Slice 300µm. 0.65 V vs. SSCE. Collaboration with J Rossier, A Rancillac & B Cauli, UMR CNRS 7637, ESPCI, Paris. Unpublished results. 2004. ~ V Action Potential Firing NO° 0.1 µM NO°
    • Modeling NO° Release in Brain
    • Modeling NO° Release in Brain
    • Modeling NO° Release in Brain
  • Acknowledgements
    • Dr Stéphane ARBAULT
    • Dr Yann BOURET
    • Dr Marie ERARD
    • Dr Yuehong TONG
    • Dr Imelda BONIFAS
    • Dr Manon GUILLE
    • Dr Frédéric LEMAîTRE
    • Mr Yann VERCHIER
    • Miss Cécile CROZATIER
    • Dr Issa TAPSOBA
    • ENS:
    • Prof Jean ROSSIER
    • Dr Bruno CAULI
    • Dr Armelle RANCILLAC
    • ESPCI (Paris):
    • Prof Irina SVIR
    • Dr Alexander OLEINICK
    • Dr Oleksiy KLYMENKO
    • KNURE (Kharkov, Ukraine):