Cannabinoid Type 1 Receptors in Astrocytes. Giovanni Marsicano, PhD
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1 Cannabinoid Type 1 Receptors in Astrocytes EndoCannabinoids and NeuroAdaptation INSERM U862 NeuroCentre Magendie Bordeaux, France 1 Cannabinoid type 1 receptors in astrocytes 1964 OH O 9 -Tetrahydrocannabinol O NH Anandamide OH Learning 3000 a.c. HPA axis plasticity Pain perception Food Intake Rewarding 2 Inflammation Locomotion Sleep Neurogenesis Sexual Behaviour How do 1 receptors do all these beautiful things 3 1
2 Text book: mechanisms of action of 1 THC/AEA 1 + K + ir + MAPK - Ca 2+ camp + PKA G i/o - - adenylyl cyclase P ATP K + A 4 hyperpolarization Decrease of neuronal excitability Text book: endocannabinoids Extracellular Depolarization Release EC Uptake Membrane Precursors + AA Precursor Enzyme 2 EC Ca 2+ + camp ATP PKA + Degradation Degrading Enzymes EC Intracellular 5 Text book: Es are retrograde neurotransmitters Pre-Synaptic GABA/Glu Ca 2+ Action potential Retrograde suppression of neurotransmitter release EC 6 Depolarization EC Ca 2+ Post-Synaptic 2
3 Where is 1 expressed Many different brain regions Distinct neuronal subpopulations Local, inhibitory GABAergic network CA3 CA1 DG Principal projecting circuits Entorhinal Cortex Hippocampus Amygdala Marsicano and Lutz, Are there cell-type specific functions of 1 Conditional Knock-out The Cre/loxP system loxp Cre Sequence X 8 Conditional Knock-out The Cre/loxP system Wild type loxp Gene of interest Cre recombinase modified = "wild type" or floxed allele Tissue-specific Transgenic expression of Cre recombinase 9 Tissue-specific mutant 3
4 Deletion of 1 from specific neuronal populations Cre/LoxP system 1 floxed mice crossed with NEX-Cre or Dlx 5/6-Cre mice ISH analysis of 1 mrna expression Cortical Glutamatergic Neurons (Glu-1-KO) GABAergic Neurons (GABA-1-KO) 10 Monory et al., Neuron 2006, PLoS, 2007; Bellocchio et al., Nat Neurosci 2010 Deletion of 1 from specific neuronal populations Immunohistochemical analysis of 1 protein in conditional mutants 1-KO Glu-1-KO GABA-1-KO 11 GABAergic neurons contain the large MAJORITY of 1 in the brain Cortical glutamatergic neurons contain the MINORITY of 1 in the brain 12 Differential roles of 1 in neurons Fasting-induced food intake Fasted 1-KO eat less than controls (Di Marzo et al., Nature 2001) F.I. (grams) F.I. (grams) Glu-1-KO (Deletion in cortical glutamatergic neurons) 1 2 Time (h) (Deletion in all cells of the body) Full 1-KO 1 2 Time (h) GABA-1-KO (Deletion in GABAergic neurons) F.I. (grams) Time (h) 4
5 What about astrocytes Until 2008, there was debate on the presence of 1 in astrocytes (only in culture) 2007: We start generating GFAP-1-KO mice (hgfap-creert2 from F. Kirchhoff, Hirrlinger et al., 2006) 2008: Electrophysiological and functional evidence (Navarrete & Araque, 2008) Pedro Grandes Bilbao Astrocyte 1 GFAP-1-KO neur astroglial onal 1 13 Han et al., Cell 2012 Cannabinoids induce hippocampal LTD in vivo Anaesthetized rats Xia Zhang University of Ottawa, Canada Systemically Intra CA1 14 Cannabinoids induce hippocampal LTD in vivo Anaesthetized rats 1 R dependent Systemically 1 R antagonist AM281: 10 min before HU min after HU
6 Mechanisms of in vivo -LTD Glu GABA AP-5: selective antagonist (intra-ca1) 16 Mechanisms of in vivo -LTD Glu -mediated LTD endocytosis Collingridge et al., 2010 GABA Tat-GLUR2: endocytosis blocker 17 Mechanisms of in vivo -LTD Glu GABA 1 R 18 subtype -specific deletion of in mouse GLU- -KO GABA- -KO -LTD does not depend on neuronal 1 receptors Where is the responsible 1 R 6
7 Mechanisms of in vivo -LTD Glu Astrocyte GABA 19 astrocyte GFAP- 1 R-KO mice astroglial 1 neuronal 1 neuro nal GFAP-1-KO -LTD depends on astroglial 1 Receptors 20 What is -LTD good for Working memory Transient holding and processing of new / already stored information Important for reasoning, comprehension, learning, memory updating 30 s THC 1 Carlini et al., 1973; Ranganathan and D Souza, 2006 How does THC alter working memory What is -LTD good for A Delayed non matching to sample in the T-maze Training B 30 s A Correct choice Test B 21 7
8 What is -LTD good for Delayed matching to place in the Morris water maze Vehicle THC Session 23 X1 X+1 Start Time and distance gain from Trial 1 to Trial What is -LTD good for A Delayed non matching to sample in the T-maze Training Test Ro25,6981 and ifenprodil: NR2B-preferring antagonists Tat-GluR2: inhibitor of GluR2 endocytosis B 30 s Correct choice Cannabinoid effect on WM depends on and AMPA A B 24 What is -LTD good for Delayed matching to place in the Morris water maze Start Vehicle THC Session 12 3 X X+1 subtype -specific deletion of in mouse GLU- 1 R-KO Time and distance gain from Trial 1 to Trial 4 GABA- 1 R-KO Cannabinoid effect on WM does not depend on neuronal 1 Receptors 1 R 8
9 25 What is -LTD good for Delayed matching to place in the Morris water maze Start Vehicle THC Session 12 3 X X+1 Astrocyte GFAP- 1 R-KO mice Time and distance gain from Trial 1 to Trial 4 Astrocyte GFAP-1-KO Cannabinoids impair working memory through astroglial 1 Receptor 1 R Mechanisms of cannabinoid impairment of working memory Working Memory impairments 1 receptors mediate cannabinoid-induced in vivo LTD 1 receptors mediate cannabinoid-induced impairment of working memory 26 Han*, Kesner*, Metna-Laurent* et al., Cell Thanks to ZHANG Xia, Philip KESNER (Ottawa), HAN Jing (Xi an), Franck KIRCHHOFF (Germany), Pedro GRANDES, Nagore PUENTE (Bilbao) Mathilde Love & Peace METNA Francis Grumpy CHAOULOFF Federico Sick MASSA Giovanni Little Gio BENARD Jose Mandolin CRUZ Tifany Breakfast DESPREZ Edgar El Chino SORIA Astrid Mama CANNICH Dany Blondie VERRIER Hu- Acupuncture -BO Philipp I.D. BETHGE Peggy Betty Boop VINCENT Michele Buonarroti COLAVITA Arnau Cataluña BUSQUETS Roman I am here SERRAT Laurie Russian ROBIN Carolina MUGURUZA Merryl MALEZIEUX 9
10 28 10
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