Lecture 9 Outline PDF

2024-11-20 Molecular Basis of Synaptic Plasticity II NROC36H3F © Arruda Carvalho UTSC Lecture Outline CamKII Structure and Functions Mechanisms...

2024-11-20 Molecular Basis of Synaptic Plasticity II NROC36H3F © Arruda Carvalho UTSC Lecture Outline CamKII Structure and Functions Mechanisms Contribution to Learning and Memory CREB LTP and Synaptic Capture Excitability IEGs Definition Engrams and LTP Arc and a novel mode of transmission BDNF © Arruda © 2018 Arruda Carvalho Carvalho UTSC winterUTSC term 1  2024-11-20 Lecture Outline CamKII Structure and Functions Mechanisms Contribution to Learning and Memory CREB LTP and Synaptic Capture Excitability IEGs Definition Engrams and LTP Arc and a novel mode of transmission BDNF © Arruda © 2018 Arruda Carvalho Carvalho UTSC winterUTSC term From LTP to CaMKII Late phase of LTP in the Schaffer collateral pathway Early LTP Principles of Neural Science ©McGraw Hill © Arruda Carvalho UTSC 2  2024-11-20 CaMKII is Abundant in the Postsynaptic Density CamKII constitutes 1–2% of the brain’s total protein. It is enriched at synapses and is the main protein of the postsynaptic density Lisman, Nat Rev Neuro 2002 © Arruda Carvalho UTSC Lecture Outline CamKII Structure and Function Mechanisms Contribution to Learning and Memory CREB LTP and Synaptic Capture Excitability IEGs Definition Engrams and LTP Arc and a novel mode of transmission BDNF © Arruda © 2018 Arruda Carvalho Carvalho UTSC winterUTSC term 3  2024-11-20 CaMKII: Structure CaMKII comprises a family of 28 similar isoforms that are derived from four genes (α, β, γ and δ). The α- and β-subunits are the predominant isoforms in the brain ATP- and substrate- binding sites, sites Directs targeting to for interaction with specific intracellular Acts as a ‘gate’ that anchoring proteins sites and modifies binds to the catalytic domain and inhibits sensitivity to enzyme activity Ca2+/calmodulin. The association domain allows for the formation of a non-dissociable holoenzyme of 12 subunits Lisman, Nat Rev Neuro 2002 © Arruda Carvalho UTSC CaMKII: Structure The catalytic/regulatory domains of the 12 subunits form two hexameric rings. Each of these domains is linked through a narrow stalk to the central mass of the molecule. This mass is composed of the 12 association domains, which are assembled into a gear- shaped structure that is formed by six slanted flanges. Hell, Neuron 2014 Lisman, Nat Rev Neuro 2002 © Arruda Carvalho UTSC 4  2024-11-20 CaMKII: Structure and Function There is a region within the autoinhibitory domain that resembles protein substrates. This pseudosubstrate region binds to the catalytic domain at the substrate- binding site (S site), inhibiting the enzyme (acts as a gate) Lisman, Nat Rev Neuro 2002 © Arruda Carvalho UTSC CaMKII: Structure and Function Active Ca2+/calmodulin binds to a region that overlaps with the This exposes the Thr286 on the pseudosubstrate region, autoinhibitory domain opening the gate and thereby activating the subunit Lisman, Nat Rev Neuro 2002 © Arruda Carvalho UTSC 5  2024-11-20 CaMKII: Structure and Function Thr286 phosphorylation renders the enzyme persistently active: T-site binding is required to position the pseudosubstrate sequence so that it inhibits the S site. After Thr286 becomes phosphorylated, binding to the T site cannot occur. As a result, the autoinhibitory domain cannot inhibit the S site and the kinase remains active Lisman, Nat Rev Neuro 2002 © Arruda Carvalho UTSC CaMKII: Structure and Function This opening of the gate can be achieved through: 1. Phosphorylation of Thr286 site by a neighbouring subunit, or 2. Binding of T side 2 1 to the NMDAR NR2B subunit. Either is sufficient to keep the gate open and the enzyme active even after dissociation of Calmodulin Lisman, Nat Rev Neuro 2002 © Arruda Carvalho UTSC 6  2024-11-20 CaMKII: Activation States Brief activation: No autophosphorylation. Calmodulin dissociates within less than 1 s after Ca2+ levels fall Short-term persistent activation: autophosphorylation of thr286. Activity persists after Ca2+ falls, but declines if Thr286 becomes dephosphorylated Lisman, Nat Rev Neuro 2002 Long-term persistent activation: After LTP. When the rate of autophosphorylation exceeds that of dephosphorylation. This is achieved when a threshold number of kinase sites are phosphorylated, working as a switch. The ‘on’ state of the switch can last very long, because the kinase acts faster than the PSD phosphatase on Thr286 sites. © Arruda Carvalho UTSC Lecture Outline CamKII Structure and Functions Mechanisms Contribution to Learning and Memory CREB LTP and Synaptic Capture Excitability IEGs Definition Engrams and LTP Arc and a novel mode of transmission BDNF © Arruda © 2018 Arruda Carvalho Carvalho UTSC winterUTSC term 7  2024-11-20 LTP Drives CamKII persistent Activity Lisman, Nat Rev Neuro 2002 © Arruda Carvalho UTSC LTP Requires CamKII Persistent Activity Lisman, Nat Rev Neuro 2002 © Arruda Carvalho UTSC 8  2024-11-20 CamKII and LTP Enhance Synaptic Transmission through the Same Mechanism Perfusion of active CaMKII on CA1 pyramidal cells Increase in amplitude of spontaneous synaptic potentials Lisman, Nat Rev Neuro 2002; From Lledo et al., PNAS 1995 © Arruda Carvalho UTSC CamKII and LTP Enhance Synaptic Transmission through the Same Mechanism No LTP Perfusion of active CaMKII LTP Increased transmission in the non-LTP group suggests occlusion between CaMKII-induced potentiation and LTP induction Lisman, Nat Rev Neuro 2002; From Lledo et al., PNAS 1995 © Arruda Carvalho UTSC 9  2024-11-20 CamKII: Mechanisms to Impact Transmission 1. Association with NMDARs (predominantly NR2B, but also NR1), which acts as a catalyst for autophosphorilation of CamKII and keeps CaMKII in the PSD Lisman, Nat Rev Neuro 2002 © Arruda Carvalho UTSC CamKII: Mechanisms to Impact Transmission Under resting conditions, F-actin binds to the -subunit of CaMKII and holds it away from synapses. Ca2+ elevation causes the kinase to dissociate from actin and move to the synapse Activation, regardless of autophosphorylation, increases the association of  and -CaMKII with the cytoplasmic c-terminal of NR2B After CAMKII binds to NR2B, it remains active even after the dissociation of Ca2+/calmodulin. This is due to a domain of NR2B being similar to the autoinhibitory domain that binds to the T site. NR2B binds to the T site, acting as a ‘wedge’ in the autoinhibitory gate, keeping it open even after dissociation of Ca2+/calmodulin. This activation (1) cannot be reversed by phosphatase activity, (2) prevents a secondary autophosphorylation of the calmodulin-binding domain (Thr305) which speeds dissociation of the kinase from synaptic sites, and (3) facilitates further autophosphorylation around the ring. Lisman, Nat Rev Neuro 2002 © Arruda Carvalho UTSC 10  2024-11-20 CamKII: Mechanisms to Impact Transmission 2. CaMKII phosphorylates AMPA receptors already at the synapse, enhancing their conductance CAMKII phosphorylates GluR1 at Ser831, increasing channel conductance Lisman, Nat Rev Neuro 2002 © Arruda Carvalho UTSC From LTP to CaMKII Recap Early LTP Principles of Neural Science ©McGraw Hill © Arruda Carvalho UTSC 11  2024-11-20 CamKII: Mechanisms to Impact Transmission 3. Regulation of trafficking of AMPARs Lisman, Nat Rev Neuro 2002 © Arruda Carvalho UTSC CamKII: Mechanisms to Impact Transmission Recap 3.1 Regulation of trafficking of AMPARs: SAP97/Myosin VI Synapse-associated protein 97 (SAP97) binds to the GluA1 PDZ domain. Following CaMKII phosphorylation, SAP97-Myosin VI complex delivers those GluA1-containing AMPARs to synapses Collingridge et al., Nat Rev Neuro 2004 © Arruda Carvalho UTSC 12  2024-11-20 CamKII: Mechanisms to Impact Transmission 3.2 Regulation of trafficking of AMPARs: Stargazin CamKII-mediated phosphorylation of Stargazin enables its binding to PSD95, which allows for stabilization of AMPARs in the synapse Mod from Lisman et al., Nat Rev Neuro 2012 © Arruda Carvalho UTSC CamKII: Mechanisms to Impact Transmission 3.2 Regulation of trafficking of AMPARs: Stargazin CamKII and Stargazin In control CA1 pyramidal cells or cells expressing wild-type stargazin, LTP can be induced (arrow denotes start of LTP induction protocol). However, LTP is blocked in CA1 pyramidal cells expressing a mutant variant of stargazin in which the nine potential CaMKII or protein kinase C serine phosphorylation sites are mutated to alanine residues (S9A). Lisman et al., Nat Rev Neuro 2012 © Arruda Carvalho UTSC 13  2024-11-20 CamKII: Mechanisms to Impact Transmission 2-3. Regulation of function and trafficking of AMPARs Mod from Lisman et al., Nat Rev Neuro 2012 © Arruda Carvalho UTSC CamKII: Mechanisms to Impact Transmission 4. Regulation of spine size through CaMKII-F-actin interaction CaMKII stabilizes F-actin structures. Activity-dependent binding of CamKII to NR2B allows for F-actin re-structuring and changes in spine size Shonesy et al., Prog Mol Biol Transl Sci. 2014 © Arruda Carvalho UTSC 14  2024-11-20 CamKII: Mechanisms to Impact Transmission Synapse-specificity After LTP induction by local glutamate uncaging (arrow), transient activation of CaMKII can be observed (red denotes high activation) Stimulation of a spine (arrow) can cause a persistent increase in spine volume and translocation of CaMKII to spines (red denotes high levels of this kinase) Lisman et al., Nat Rev Neuro 2012 © Arruda Carvalho UTSC CamKII in vivo: Synaptic Maturation During early development, retinotectal synapses are primarily silent, lacking AMPA receptors. At this time, CaMKII is expressed at low levels. Increasing CAMKII expression in this preparation leads to unsilencing, similar to LTP induction. Active CaMKII (red bars) increases the AMPA-, but not the NMDA-receptor mediated component of the synaptic response compared with uninfected controls (green bars) or with controls infected with β-galactosidase (blue bars). Lisman, Nat Rev Neuro 2002 © Arruda Carvalho UTSC 15  2024-11-20 Lecture Outline CamKII Structure and Functions Mechanisms Contribution to Learning and Memory CREB LTP and Synaptic Capture Excitability IEGs Definition Engrams and LTP Arc and a novel mode of transmission BDNF © Arruda © 2018 Arruda Carvalho Carvalho UTSC winterUTSC term CamKII in vivo: Learning and Memory Silva et al., Science, 1992a © Arruda Carvalho UTSC 16  2024-11-20 CamKII in vivo: Learning and Memory Silva et al., Science, 1992b © Arruda Carvalho UTSC CamKII in vivo: Learning and Memory A point mutation was introduced into the aCaMKII gene that blocked the autophosphorylation of threonine at Probe test position 286 (Thr286) WT aCaMKIIT286A-129B6F2 mutants Giese et al., Science, 1998 © Arruda Carvalho UTSC 17  2024-11-20 CamKII and LTP CamKII inhibitors block early and late phase LTP, indicating cAMP can only strengthen synaptic transmission via a CaMKII dependent mechanism. © Arruda Carvalho UTSC Lecture Outline CamKII Structure and Functions Mechanisms Contribution to Learning and Memory CREB LTP and Synaptic Capture Excitability IEGs Definition Engrams and LTP Arc and a novel mode of transmission BDNF © Arruda © 2018 Arruda Carvalho Carvalho UTSC winterUTSC term 18  2024-11-20 Mice with Disrupted CREB Activity are Impaired in Long but not Short-term Fear Conditioning Recap  and  isoforms 30min 24h Bourtchuladze et al., Cell 1994 © Arruda Carvalho UTSC CREB Inhibition Leads to Long- but not Short-Term Memory Deficits The mammalian genome contains three CREB-related genes—CREB, CREM, and ATF-1— all of which are heavily spliced, generating multiple gene-regulating complexes Transgenic mice expressing KCREB, a mutant of human CREB that is a potent dominant- negative inhibitor and inhibits all three CREB family transcription factors in CA1 © Arruda Carvalho UTSC 19  2024-11-20 CREB Inhibition Leads to Long- but not Short-Term Memory Deficits Novel object recognition 1h post-training 24h post-training Pittenger et al., Neuron, 2002 © Arruda Carvalho UTSC Lecture Outline CamKII Structure and Functions Mechanisms Contribution to Learning and Memory CREB LTP and Synaptic Capture Excitability IEGs Definition Engrams and LTP Arc and a novel mode of transmission BDNF © Arruda © 2018 Arruda Carvalho Carvalho UTSC winterUTSC term 20  2024-11-20 CREB Inhibition Impairs some Forms of LTP, but not Others Forskolin Strong Theta burst Pittenger et al., Neuron, 2002 control dCA1-KCREB Some experimental forms of plasticity may bypass the requirement for CREB © Arruda Carvalho UTSC Increased CREB Activity Leads to Increased Expression of CREB-Dependent Genes Barco et al., Cell, 2002 tTA system to express VP16-CREB (VC), a constitutively active form of CREB, in forebrain neurons © Arruda Carvalho UTSC 21  2024-11-20 Increased CREB Activity Facilitates L-LTP A single 100 Hz train (1 s) evoked E-LTP that lasts up to 2 hr in wild-type animals (white bars), but L- LTP lasting more than 6 hr in VP16- CREB mice (black bars) Lower threshold to induce L-LTP Barco et al., Cell, 2002 tTA system to express VP16-CREB, a constitutively active form of CREB, in forebrain neurons © Arruda Carvalho UTSC CREB Overexpression Increases Long-Term Memory Josselyn et al., J Neurosci, 2001 © Arruda Carvalho UTSC 22  2024-11-20 CREB is Phosphorylated Following Fear Training Han et al., Science, 2007 © Arruda Carvalho UTSC LA Neurons with Increased CREB are more Likely to be Recruited into the Memory Trace Han et al., Science, 2007 © Arruda Carvalho UTSC 23  2024-11-20 Ablation of LA Neurons Overexpressing CREB Blocks Memory Expression Han et al., Science, 2009 © Arruda Carvalho UTSC Lecture Outline CamKII Structure and Functions Mechanisms Contribution to Learning and Memory CREB LTP and Synaptic Capture Excitability IEGs Definition Engrams and LTP Arc and a novel mode of transmission BDNF © Arruda © 2018 Arruda Carvalho Carvalho UTSC winterUTSC term 24  2024-11-20 CREB Overexpressing Neurons Are More Excitable Yiu et al., Neuron 2014 © Arruda Carvalho UTSC Increasing Intrinsic Excitability Mimics CREB-Mediated Potentiation of Memory How Could CREB increase excitability? Through which mechanism? (disrupts function of the voltage- dependent K+ channel KCNQ2, increasing neuronal excitability) Yiu et al., Neuron 2014 © Arruda Carvalho UTSC 25  2024-11-20 Potential Mediators of CREB Effect on Excitability Zhang et al., PNAS 2005 © Arruda Carvalho UTSC Lecture Outline CamKII Structure and Functions Mechanisms Contribution to Learning and Memory CREB LTP and Synaptic Capture Excitability IEGs Definition Engrams and LTP Arc and a novel mode of transmission BDNF © Arruda © 2018 Arruda Carvalho Carvalho UTSC winterUTSC term 26  2024-11-20 Immediate Early Genes (IEGs) Rapid and transient induction by pre-existing transcription factors (without the need for de novo protein synthesis) in response to extracellular signals such as growth factors and neurotransmitters Control Post-seizure Relatively short half-life Arc, c-fos, Homer1a, zif268 c-fos IHC Morgan et al., Science, 1987 © Arruda Carvalho UTSC Pattern of Activity that Triggers LTP Leads to IEG Expression Abraham et al., Mol Neurobiol 1992 IEG regulatory functions are believed to trigger cascades of activity-dependent neuronal gene expression that can lead to plastic events that may be critical for memory consolidation © Arruda Carvalho UTSC 27  2024-11-20 Lecture Outline CamKII Structure and Functions Mechanisms Contribution to Learning and Memory CREB LTP and Synaptic Capture Excitability IEGs Definition Engrams and LTP Arc and a novel mode of transmission BDNF © Arruda © 2018 Arruda Carvalho Carvalho UTSC winterUTSC term Engrams and the Locus of Memory Richard Semon, Karl Lashley, 1859-1918 1890-1958 © Arruda Carvalho UTSC 28  2024-11-20 Engrams and the Locus of Memory https://soreike.wordpress.com/2013/07/19/ponder-of-the-night-eternal-sunshine-of-the-spotless-mind/ https://www.thoughtco.com/jim-carrey-on-eternal-sunshine-of-the-spotless-mind- 2419275 © Arruda Carvalho UTSC Reactivation of IEG-Expressing Neurons Triggers Memory Recall Liu et al., Nature 2012 © Arruda Carvalho UTSC 29  2024-11-20 How do IEGs Contribute to Long-Term Plasticity? How do these IEGs contribute to encoding/storing information at the individual cell level? Can we use this principle to understand how memory is encoded at a population level? NROD61: Emotional Learning © Arruda Carvalho UTSC How do IEGs Contribute to Long-Term Plasticity? c-fos KO impairs structural plasticity (sprouting of mossy fiber axons – measured by the histological method timm) in Hippocampus following seizures (kindled) Watanabe et al., J Neurosci 1996 © Arruda Carvalho UTSC 30  2024-11-20 Lecture Outline CamKII Structure and Functions Mechanisms Contribution to Learning and Memory CREB LTP and Synaptic Capture Excitability IEGs Definition Engrams and LTP Arc and a novel mode of transmission BDNF © Arruda © 2018 Arruda Carvalho Carvalho UTSC winterUTSC term How do IEGs Contribute to Long-Term Plasticity? Arc (activity-regulated cytoskeleton associated protein, aka Arg 3.1) Lyford et al., Neuron 1995 Arc is present in dendrites and soma, is induced after strong cellular activity and co-precipitates with F-actin © Arruda Carvalho UTSC 31  2024-11-20 Arc mRNA Moves to Activated Dendritic Domains Following Stimulation Arc mRNA travels very rapidly throughout the dendrites (~300μm/hr). This is independent of protein synthesis, but dependent on NMDA Steward et al., Neuron 1998 © Arruda Carvalho UTSC Arc mRNA Moves to Activated Dendritic Domains Following Stimulation Steward et al., Neuron 1998 © Arruda Carvalho UTSC 32  2024-11-20 Inhibition of Activity-dependent Arc Expression Impairs LTP and Long-Term Memory Water maze Guzowski et al., J Neurosci 2000 © Arruda Carvalho UTSC Arc Regulates AMPAR Trafficking Arc-endocytosis pathway regulates basal AMPAR levels: Arc KO neurons show reduced AMPAR endocytosis and increased steady state surface levels Chowdhury et al., Neuron 2006 © Arruda Carvalho UTSC 33  2024-11-20 Arc Regulates AMPAR Trafficking Lacking Dynamin- binding domain Arc protein interacts with dynamin to accelerate AMPA receptor endocytosis, reducing its surface expression Chowdhury et al., Neuron 2006 © Arruda Carvalho UTSC Molecular basis of LTD: mGluR-dependent Recap Collingridge et al., Nat Rev Neuro 2010 d. mGluR-LTD has been shown to require rapid (in a few minutes) local de novo protein synthesis, which depends on the PI3K-Akt-mTOR pathway. An important protein being translated is Arc, thought to help initiate dynamin-dependent endocytosis of AMPARs. © Arruda Carvalho UTSC 34  2024-11-20 Transfer of Arc mRNA Between Neurons: A Novel Signaling Pathway in the CNS? Arc self-assembles into virus-like capsids that encapsulate RNA. Endogenous Arc is released from neurons in extracellular vesicles that mediate the transfer of Arc mRNA into new target cells, where it can undergo activity-dependent translation! Parrish and Tomonaga, Cell 2018 comment on Pastuzyn et al., Cell 2018 © Arruda Carvalho UTSC Transfer of Arc mRNA Between Neurons: A Novel Signaling Pathway in the CNS? Arc encodes a protein that forms virus-like capsids Arc protein exhibits similar biochemical properties as retroviral Gag proteins Endogenous Arc protein is released from neurons in extracellular vesicles (EVs) Arc EVs and capsids can mediate intercellular transfer of Arc mRNA in neurons “We favor the idea that released Arc functions to carry intercellular cargo that alters the state of neighboring cells required for cellular consolidation of information.” Pastuzyn et al., Cell 2018 © Arruda Carvalho UTSC 35  2024-11-20 Lecture Outline CamKII Structure and Functions Mechanisms Contribution to Learning and Memory CREB LTP and Synaptic Capture Excitability IEGs Definition Engrams and LTP Arc and a novel mode of transmission BDNF © Arruda © 2018 Arruda Carvalho Carvalho UTSC winterUTSC term BDNF Neurotrophins such as brain-derived neurotrophic factor (BDNF), nerve growth factor (NGF) and neurotrophins 3 and 4 (NT3 and NT4), are regulatory factors implicated in cell development, survival, repair and plasticity During development, BDNF regulates neuronal proliferation, neuronal migration, axon path finding, dendritic growth and synapse formation BDNF binds to Tropomyosin receptor kinase B receptor (TrkB), a receptor tyrosine kinase https://www.elisagenie.com/2018/02/06/bdnf-inflammation-neuropathologies/ © Arruda Carvalho UTSC 36  2024-11-20 BDNF can Induce LTP Bramham et al., Prog Neurobiol 2005 © Arruda Carvalho UTSC BDNF is Necessary for Spatial Learning Mu et al., Brain Res 1999 © Arruda Carvalho UTSC 37  2024-11-20 BDNF induces the synthesis of PKM © Arruda Carvalho UTSC LTP Maintenance: Recap PKM PKMζ is an isoform of protein kinase C which lacks a regulatory domain and is thus constitutively active Levels of PKMζ in the hippocampus are normally low. Tetanic stimulation leads to an increase in translation of PKMζ mRNA. This mRNA is present in the CA1 neuron dendrites, enabling its local translation to rapidly alter synaptic strength through enhancing AMPAR membrane insertion Principles of Neural Science ©McGraw Hill © Arruda Carvalho UTSC 38  2024-11-20 Recap …and CamKII Protein Synthesis in Mechanically Isolated Dendrites Protein synthesis reporter in which the coding sequence of green fluorescent protein is flanked by the 5' and 3' untranslated regions from CAMKII-alpha BDNF treatment in transected neuron Aakalu et al., Neuron 2001 © Arruda Carvalho UTSC Park and Poo, Nat Rev Neuro 2013 © Arruda Carvalho UTSC 39  2024-11-20 Lecture Outline CamKII Structure and Functions Mechanisms Contribution to Learning and Memory CREB LTP and Synaptic Capture Excitability IEGs Definition Engrams and LTP Arc and a novel mode of transmission BDNF © Arruda © 2018 Arruda Carvalho Carvalho UTSC winterUTSC term 40