2011.10.14.. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 1 Development of Complex Curricula for Molecular Bionics and Infobionics Programs within a consortial* framework** Consortium leader PETER PAZMANY CATHOLIC UNIVERSITY Consortium members SEMMELWEIS UNIVERSITY, DIALOG CAMPUS PUBLISHER The Project has been realised with the support of the European Union and has been co-financed by the European Social Fund *** **Molekuláris bionika és Infobionika Szakok tananyagának komplex fejlesztése konzorciumi keretben ***A projekt az Európai Unió támogatásával, az Európai Szociális Alap társfinanszírozásával valósul meg. PETER PAZMANY CATHOLIC UNIVERSITY SEMMELWEIS UNIVERSITY sote_logo.jpg dk_fejlec.gif INFOBLOKK 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 2 Peter Pazmany Catholic University Faculty of Information Technology BEVEZETÉS A FUNKCIONÁLIS NEUROBIOLÓGIÁBA INTRODUCTION TO FUNCTIONAL NEUROBIOLOGY www.itk.ppke.hu By Imre Kalló Contributed by: Tamás Freund, Zsolt Liposits, Zoltán Nusser, László Acsády, Szabolcs Káli, József Haller, Zsófia Maglóczky, Nórbert Hájos, Emilia Madarász, György Karmos, Miklós Palkovits, Anita Kamondi, Lóránd Erőss, Róbert Gábriel, Zoltán Kisvárday, Zoltán Vidnyánszky Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 3 www.itk.ppke.hu ElectrophysiologyImre Kalló & Norbert HájosPázmány Péter Catholic University, Faculty of Information Technology I. Membrane and action potentials in neurons. II. Signal transmission at the synapses. III. Synaptic plasticity IV. In vitro and in vivo recording techniques V. Firing properties of different neuronal phenotypes. Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 4 www.itk.ppke.hu Ion concentrations and Equilibrium Potentials Distributionofionsonoppositesidesofacellularmembraneisunequalduethesemipermeabilityofthemembraneretainingthenegativelychargedproteinmoleculeswithinthecellandtheregulatedactionofionchannelsandionpumpstransferringionsfromonesideofthemembranetotheother.Thevoltagedifferencegeneratedbythealteredionicconcentrationsontheoppositesidesofthecellularmembraneiscalledthemembranepotential.Theconcentrationofsodium(Na+;145mMvs.18mM)andchloride(Cl–,120mMvs7mM)ionsarehighintheextracellularregion,whereaspotassium(K+,135mMvs.3mM)ions,alongwithlargeproteinanionshavehighconcentrationsintheintracellularregion.Calciumionsarekeptintracellularlyatnanomolarconcentrations(100nM),elevationsofwhichfromtheextracellularspace(1.2mM)andintracellularstoreschangethemembranepotential,aswellasactivatecalcium-dependentintracellularprocesses.Eachoftheionsarecharacterizedbyamembranepotentialatwhichitsflowfromonesideofthemembranetotheotherisinequilibrium,consequentlyitsnet(overall)flowiszero. Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 5 www.itk.ppke.hu Ion concentrations and Equilibrium Potentials figl Inside (mM) Outside (mM) Equilibrium potential (mM) Squid giant axon Na+ 50 440 +55 K+ 400 20 -76 Cl- 40 560 -66 Ca++ 0.4 µM 10 +145 Mammalian neuron Na+ 18 145 +56 K+ 135 3 -102 Cl- 7 120 -76 Ca++ 0.1 µM 1.2 +125 Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 6 www.itk.ppke.hu The Nernst Equation Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 7 www.itk.ppke.hu The Goldman Equation Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 8 www.itk.ppke.hu The Action Potential A. L. Hodgkin A.F. Huxley Sir J. C. Eccles The Nobel Prize in Physiology or Medicine 1963 “for their discoveries concerning the ionic mechanisms involved in excitation and inhibition in the peripheral and central portions of the nerve cell membrane" squid Squid Eccles.jpg Hodgkin.jpg Huxley.jpg Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 9 www.itk.ppke.hu What is happening during Action Potential? ActionPotential1.jpg ActionPotential2.jpg Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 10 www.itk.ppke.hu The Hodgkin-Huxley equations Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 11 www.itk.ppke.hu The Hodgkin-Huxley equations Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 12 www.itk.ppke.hu Potential changes during Action Potential squid A. L. Hodgkin A.F. Huxley ActionPotential4.jpg ActionPotential5.jpg Hodgkin.jpg Huxley.jpg Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 13 www.itk.ppke.hu Voltage clamp3.jpg Voltage clamp4.jpg The voltage clamp is a current generator The voltage clamp operates under a negative feedback mechanism Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 14 www.itk.ppke.hu Demonstration of ionic movements during Action Potential (mV) -50 -60 0 0 Outward Inward Vm Im Ic Ic Il Ic Il IK INa Current measured in the presence of TTX Current measured in the presence of TEA Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 15 www.itk.ppke.hu channel Bert Sakmann Erwin Neher The Nobel Prize in Physiology or Medicine 1991 "for their discoveries concerning the function of single ionchannels in cells" Patch Clamp.jpg Patch-clamp recording erwin_neher.jpg bert%20sakmann.jpg Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 16 www.itk.ppke.hu Direct examination of channel proteins FFig6 SingleChannel1.jpg FFig5 SingleChannel2.jpg Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 17 www.itk.ppke.hu Movement of ions during Action Potential ActionpotentialIons1.jpg Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 18 www.itk.ppke.hu Movement of ions during Action Potential ActionpotentialIons2.jpg Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 19 www.itk.ppke.hu Opening of ion channels during Action Potential figb Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 20 www.itk.ppke.hu Structure and function of voltage-gated ion channels figr figq figp Introduction to functional neurobiology: Electrophysiology FiringActivity4.jpg 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 21 www.itk.ppke.hu Great variety of firing activity of neurons FiringActivity.jpg FiringActivity1.jpg FiringActivity2.jpg FiringActivity3.jpg FiringActivity5.jpg FiringActivity6.jpg Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 22 www.itk.ppke.hu Morphology of neurons PyramidalCells.jpg Axon Dendritic tree Cell body Axon initial segment Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 23 www.itk.ppke.hu Synaptic potentials, passive and active dendrites Ref:Williams SR, Stuart GJ. Role of dendritic synapse location in the control olfaction potential output. Trends Neurosci. 2003 Mar;26(3):147-54. Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 24 www.itk.ppke.hu The asynchronous excitatory input is summed linearly, whereas the synchronous input supralinearly in the dendrites Ref:LosonczyA, Magee JC. Integrative properties of radial oblique dendrites inhippocampal CA1 pyramidal neurons. Neuron. 2006 Apr 20;50(2):291-307. Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 25 www.itk.ppke.hu The amplitude of the retrograde propagating action potential in the dendrite is dependent on the firing pattern Ref:StuartG,SakmannB.AmplificationofEPSPsbyaxosomaticsodiumchannelsinneocorticalpyramidalneurons.Neuron.1995Nov;15(5):1065-76. Ref:WilliamsSR,StuartGJ.Mechanismsandconsequencesofactionpotentialburstfiringinratneocorticalpyramidalneurons.JPhysiol.1999Dec1;521Pt2:467-82. Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 26 www.itk.ppke.hu Removal of K+ions from the extracellular space figv Kalium.jpg Glia.jpg Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 27 www.itk.ppke.hu Synaptic neurotransmission The presynaptic side Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 28 www.itk.ppke.hu Mechanism of neurotransmission FFigz Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 29 www.itk.ppke.hu Ultrastructure of axon terminals AxonTerminals1.jpg AxonTerminals3.jpg AxonTerminals2.jpg Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 30 www.itk.ppke.hu Proving the chemical nature of neurotransmission fig6 Experiment of Otto Loewi in 1920 Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 31 www.itk.ppke.hu Synaptic vesicle and its associated proteins fig5 Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 32 www.itk.ppke.hu Release of the content of synaptic vesicle FFigb FFigc FFiga Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 33 www.itk.ppke.hu Exocytosis increase the capacitance of the cell FFigd Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 34 www.itk.ppke.hu Synaptic neurotransmission The postsynaptic side Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 35 www.itk.ppke.hu Mechanism of neurotransmission FFigz Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 36 www.itk.ppke.hu Structure of postsynaptic ion channels and receptor proteins Indirect gating IonChannels1.jpg IonChannels2.jpg IonChannels3.jpg GPCR.jpg Direct gating Ionotropic receptor G protein-coupled receptor Receptor tyrosine kinase Structure of metabotropic receptors Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 37 www.itk.ppke.hu Function of channel proteins Receptors1.jpg Receptors2.jpg Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 38 www.itk.ppke.hu Fast (10ms) –ionotropic -neurotransmission Slow (100ms) –metabotropic -neurotransmission Excitatory: glutamate receptors -AMPA r. (Na+andK+) kainater. (Na+andK+) NMDA r. (Na+, K+ andCa2+) acetylcholine receptors -nicotine r. (Na+, K+ andCa2+) serotonin receptors -5HT3 r.(Na+, K+ andCa2+) Inhibitory: GABA receptors -GABAar. (Cl-) glycine receptors (Cl-) glutamate receptors -mGluRr. (mGluR1-8) GABA receptors -GABAbr. acetylcholine receptors -muscarinicr. (M1-5) serotonin receptors -5HT1-8 r. dopamine receptors -D1-D6 r. adrenergic receptors -alpha 1,2 r.; beta1,2 r. histamine -H1-3 r. Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 39 www.itk.ppke.hu EPSP Excitatory neurotransmission (e.g. glutamate or acetylcholine receptors) Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 40 www.itk.ppke.hu IPSP Inhibitory neurotransmission (e.g. GABAa receptors) Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 41 www.itk.ppke.hu Structure and modifiability of a large variety of postsynaptic receptors. PostsynapticR1.jpg PostsynapticR2.jpg Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 42 www.itk.ppke.hu Cholinergic pathways in the rat brain fig1 Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 43 www.itk.ppke.hu Serotonergic pathways in the rat brain fig2 Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 44 www.itk.ppke.hu Noradrenergic pathways in the rat brain Pathway1.jpg Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 45 www.itk.ppke.hu Dopaminergic pathways in the rat brain Pathway2.jpg Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 46 www.itk.ppke.hu Activation of second messengers I. FFigs Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 47 www.itk.ppke.hu Retrograde signalization i) Gaseous: nitrogen monoxide (NO), carbon monoxide (CO) ii) Peptides: BDNF, dynorphin iii) Lipids: endocannabinoids, arachidonyl acid iv) Classical neurotransmitters: GABA, glutamát Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 48 www.itk.ppke.hu Retrograde synaptic signalization Inhibitory postsynaptic potentials -IPSP Ref:Wilson RI, NicollRA. Endocannabinoid signaling in the brain. Science. 2002Apr 26;296(5568):678-82. Review. Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 49 www.itk.ppke.hu Retrograde synaptic signalization Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 50 www.itk.ppke.hu Short term plasticity of synapses It is dependent on the quality of target elements It can be depressive, facilitative and temporally stabile Causes of depression: i) probability of transmitter release is high ii) desensitation of receptors iii) intracellular factors (e.g. spermin) Causes of facilitation: accumulation of Ca2+ in the presynaptic terminal It can be influenced by e.g. activation of presynaptic receptors Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 51 www.itk.ppke.hu Elements of the neocortical network PC B M Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 52 www.itk.ppke.hu Elements of the neocortical network The somatostatin-containing cells showbitufted(B)morphology, which innervate the dendritic tree of other neurons. The parvalbumin-containing cells show multipolar(M) morphology,which innervate the perisomaticregion of other neurons(basketoraxo-axoniccells) Ref:Reyes A, Lujan R, RozovA, BurnashevN, SomogyiP, SakmannB. Target-cell-specific facilitation and depression in neocortical circuits. Nat Neurosci. 1998 Aug;1(4):279-85. Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 53 www.itk.ppke.hu Spatially and temporally distinguished role of multipolar perisomatic and bitufted dendritic inhibitory cells: The functional difference originate from the different plasticity of excitatory synapses Ref:PouilleF, ScanzianiM. Routing of spike series by dynamic circuits in thehippocampus. Nature. 2004 Jun 17;429(6993):717-23. Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 54 www.itk.ppke.hu Spatially and temporally distinguished role of multipolar perisomatic and bitufted dendritic inhibitory cells: The functional difference originate from the different plasticity of excitatory synapses Ref:PouilleF, ScanzianiM. Routing of spike series by dynamic circuits in thehippocampus. Nature. 2004 Jun 17;429(6993):717-23. Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 55 www.itk.ppke.hu Long-term plasticity of synapses It is dependent on the quality of traget elements It can be LTP –Long Term Potentiation or LTD –Long Term Depression Methods which can induce: i) high frequency stimulus of the fibers (Bliss & Lomo, 1973) ii) synchronous co-activation of pre-and postsynaptic cells (plasticity window: presynaptic and postsynaptic spikeslessthan 15 ms apart) iii) temporal difference between the activation of pre-and postsynaptic cells Mechanisms: NMDA-dependent, non-NMDA dependent, receptor insertion, receptor subunit exchange, retrograde messengers etc. Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 56 www.itk.ppke.hu Spike-time dependent plasticity -STDP Ref:MarkramH, LübkeJ, FrotscherM, SakmannB. Regulation of synaptic efficacy by coincidence of postsynaptic APs and EPSPs. Science. 1997 Jan 10;275(5297):213-5. Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 57 www.itk.ppke.hu Spike-time dependent plasticity -STDP Ref:MarkramH, LübkeJ, FrotscherM, SakmannB. Regulation of synaptic efficacy by coincidence of postsynaptic APs and EPSPs. Science. 1997 Jan 10;275(5297):213-5. Introduction to functional neurobiology: Electrophysiology 2011.10.14. TÁMOP –4.1.2-08/2/A/KMR-2009-0006 58 www.itk.ppke.hu Direct activation of astroglial cells through depolarization or Ca2+ uncaging can enhance GABA release via kainate receptors Ref:Kang J, Jiang L, Goldman SA, NedergaardM. Astrocyte-mediated potentiationof inhibitory synaptic transmission. Nat Neurosci. 1998 Dec;1(8):683-92.