Control of dopamine ascending pathways by central 5-HT system: implications for treatment of Parkinson’s disease Umberto Spampinato Bordeaux 2 University – INSERM U862 Bordeaux - France Summer School of Neuroscience (5) July, 7 -13, 2007, Catania, Italy

5-HT/DA interaction: anatomical basis innervates DA ascending pathways Na+/K+ channel 5-HT1A 5-HT1B AMPc 5-HT4 5-HT6 5-HT7 5-HT2A 5-HT2C IP3/DAG highly represented in DA regions Frontal cortex n. Accumbens DA 5-HT striatum DA 5-HT 5-HT 5-HT DA DA VTA / SN DR innervates DA ascending pathways  DA neuron activity

5-HT/DA interaction and neuropsychiatric disorders Na+/K+ channel 5-HT1A 5-HT1B AMPc 5-HT4 5-HT6 5-HT7 5-HT2A 5-HT2C IP3/DAG Schizophrenia Depression Drug Addiction Parkinson’s disease Frontal cortex n. Accumbens DA 5-HT striatum DA 5-HT 5-HT 5-HT DA DA VTA / SN DR target for improved treatment of DA related diseases

5-HT system and Parkinson’s disease Preclinical data (rat): central 5-HT transmission Catalepsy central 5-HT transmission (SSRI) Clinical data: Park symptoms dyskinesia central 5-HT transmission (SSRI) SSRI: recent preclinical (MPTP monkey) and clinical studies: pas Δ L-dopa-induced dyskinesia / park symptoms

5-HT/DA interaction and neuropsychiatric disorders Na+/K+ channel 5-HT1A 5-HT1B AMPc 5-HT4 5-HT6 5-HT7 5-HT2A 5-HT2C IP3/DAG Schizophrenia Depression Drug Addiction Parkinson’s disease Frontal cortex n. Accumbens DA 5-HT striatum DA 5-HT 5-HT 5-HT DA DA VTA / SN DR selective action on 5-HT receptor subtypes

5-HT1 receptors and Parkinson’s disease LID MPTP monkey Park patients 5-HT1AR agonists 8-OH-DPAT sarizotan (low doses) buspirone ↑ duration of L-dopa action 5-HT1BR agonists MDMA (ecstasy): 5-HT1A/ 1B stim ( )

5-HT2 receptors and Parkinson’s disease 5-HT2C antagonism is effective in animal models of Parkinson’s disease 5-HT2A/2C antagonism catalepsy anti-parkinsonian action DA agonists L-dopa induced dyskinesia (MPTP primates) 5-HT2C receptor binding is increased in the SN ret of parkinsonian patients - atypical antipsychotic (clozapine) - antidepressant mirtazapine L-dopa-induced psychosis, LID / tremor tremor - LID 5-HT2A/2C antagonism may participate to the therapeutic benefit of: mechanisms ?

key factor for the 5-HT-DA interaction 5-HT2C receptor 5-HT3 Na+/K+ channel 5-HT1A 5-HT1B AMPc 5-HT4 5-HT6 5-HT7 5-HT2A 5-HT2C IP3/DAG Schizophrenia Depression Drug addiction Parkinson’s disease Frontal cortex n. Accumbens DA 5-HT striatum DA 5-HT 5-HT2C 5-HT DA DA VTA / SN DR key factor for the 5-HT-DA interaction

5-HT2C receptor: cellular location indirect and direct control double-labeling in situ hybridization non-DA neurons (GABA) Eberle-Wang et al., 1997 SN GAD + 5-HT2CR mRNA positive GAD mRNA positive VTA-DA neuron subpopulation Bubar & Cunningham, 2007 TH TH + 5-HT2CR 5-HT2CR indirect and direct control of DA neuron activity

5-HT2C Rs control DA release and/or transmission electrophysiological biochemical behavioral studies Fr Cortex / NAc DA release DA transmission 5-HT2C 5-HT2C receptor control of DA pathways: VTA composite responses involving different populations of 5-HT2C receptors in multiple brain nuclei DA controlling NAc DA release 5-HT2C and/or DA transmission

5-HT2C receptor in the basal ganglia highly represented key role in controlling basal ganglia output Cortex Striatum/NAc SNc/VTA DA GPe VL NST GPi SNr RD/RM 5-HT NPP 5-HT2C Alexander et Crutcher, 1990; Obeso et coll., 2001

5-HT2C receptor: molecular and functional properties COOH TM1 TM2 TM3 TM4 TM5 TM6 TM7 NH2 G-protein coupled receptor family Molecular property constitutive activity activation of intracellular signaling in the absence of agonist stimulation - log[drug], M IP3 accumulation (% basal) 150 - 50 100 10 - 11 9 7 5 5-HT - 5HT2C agonist 5HT2C inverse agonist inverse agonist ↓ constitutive activity Le récepteur 5HT2C, est un des 14 récepteurs à la 5HT connus. Il appartient à la superfamille des récepteurs couplés à une protéine G. Ce récepteur présente des propriétés intéressantes: au niveau de ses propriétés moléculaires, il a été montré qu’il possède une activité constitutive, CAD une activation de la signalisation intracellulaire en l’absence de stimulation par l’agoniste. De plus, au niveau fonctionnel, ce récepteur exerce un contrôle inhibiteur tonique et phasique sur l’activité des voies DA centrales, dont la voie mésoaccumbale. pre-mRNA editing variety of receptor isoforms with different G protein coupling

5-HT2C receptor: molecular and functional properties COOH TM1 TM2 TM3 TM4 TM5 TM6 TM7 NH2 G-protein coupled receptor family Functional property “ 5-HT2C receptor mediates tonic inhibition of neuronal network excitability ” Le récepteur 5HT2C, est un des 14 récepteurs à la 5HT connus. Il appartient à la superfamille des récepteurs couplés à une protéine G. Ce récepteur présente des propriétés intéressantes: au niveau de ses propriétés moléculaires, il a été montré qu’il possède une activité constitutive, CAD une activation de la signalisation intracellulaire en l’absence de stimulation par l’agoniste. De plus, au niveau fonctionnel, ce récepteur exerce un contrôle inhibiteur tonique et phasique sur l’activité des voies DA centrales, dont la voie mésoaccumbale. Tecott et al., 1995 fine-tuning of DA neuron activity

Aims Basal DA neuron activity DA effects of antipsychotic drugs Role of central 5-HT2C receptors in the control of the mesoaccumbens and the nigrostriatal DA pathways Basal DA neuron activity role of the 5-HT2CR constitutive activity DA effects of antipsychotic drugs Haloperidol – Clozapine Mesoaccumbens DA pathway & VTA / NAc 5-HT2C Rs agonists versus inverse agonists effects monitoring NAc DA release by in vivo microdialysis

Experimental procedure : intracerebral microdialysis time (min) Dialysates (30 µl) HPLC-ECD Perfusion Pump (flow rate: 2 µl/min-1) microdialysis probes Striatum 1 mm NAc Striatum NAc SNc ATV

Experimental procedure Representation of the results 100 150 200 DA (% of baseline ± s.e.m.) -30 30 60 90 120 150 180 Time (min)

I. 5-HT2C receptors and basal DA release Tonic and inhibitory control on DA neuron activity Cumulative i.v. doses of mesulergine (MES) on basal DA neuron firing rate in the VTA Prisco et al., 1994 Dose-response of mesulergine (0.1/0.2/0.5 mg/kg) on DA release at terminals (NAC/Striatum) Spampinato et al., 1997 Mesulergine, a non selective 5-HT2C antagonist

I. 5-HT2C receptors and basal DA release Striatum NAC Tonic and inhibitory control selective 5-HT2C agents SNc SB 206553 5-HT2C/2B antagonist SB 242084 5-HT2C antagonist SB 243213 5-HT2C antagonist Ro 60-0175 5-HT2C agonist VTA

5-HT2C receptors and basal DA release Striatum NAC Tonic and inhibitory control 5-HT2C agents SNc VTA preferential control of the mesoaccumbens DA pathway ? Sensitivity / magnitude

Effect of 5-HT2C agents on DA release assessed by in vivo microdialysis striatum n. accumbens mg/kg 5-HT2C agonists 1 mCPP Di Giovanni et al., 2000 1 MK-212 Di Giovanni et al., 2000 n.d. 5 Willins & Meltzer, 1998 1 Ro60-0175 Di Matteo et al., 1999 2.5 Gobert et al., 2000 -40 -30 -20 -10 -10 -20 -30 -40 mg/kg 5-HT2C antagonists n.d. Di Matteo et al., 1998 1 SB 206553 n.d. 2.5 Di Matteo et al., 1998 5 Porras et al., 2002 10 Gobert et al., 2000 n.d. 2.5 Di Giovanni et al., 2000 5 Di Matteo et al., 1999 10 SB 242084 Di Matteo et al., 1999 Gobert et al., 2000 10 +80 +60 +40 +20 +20 +40 +60 +80 DA release (%)

Do 5-HT2C receptors exert a preferential control of the mesoaccumbens DA pathway ? Striatum NAC 5-HT2C agents Dose-response studies Simultaneous monitoring of DA release in both regions SNc VTA

5-HT2C receptors and basal DA release SB 206553 : 5-HT2B/2C antagonist Striatum NAC DA (% of baseline) Time (min) -30 30 60 90 120 150 180 100 200 Veh. 1 mg/kg ** 5 mg/kg *** 10 mg/kg *** 1 mg/kg **p<0.01, ***p<0.001 Fisher’s test Tonic inhibitory control on DA release in both the striatum and NAC

5-HT2B receptors and basal DA release 5-HT2B antagonist SB 204741 5-HT2B agonist BW 723C86 NAc DA (% of baseline) Vehicle BW 723C86, 1 mg/kg, ip Vehicle SB 204701, 10 mg/kg, sc NAc DA (% of control) Time (min) Gobert et al., 2000 Di Matteo et al., 2000 5-HT2B receptors: no influence on basal DA release NAC = striatum effect of SB 206553 → R 5-HT2C

5-HT2C receptors and basal DA release SB 242084 : 5-HT2C antagonist Striatum NAC * * ** ** * * *p<0.05, **p<0.01 Fisher’s test Tonic inhibitory control on DA release in both the striatum and NAC

5-HT2C receptors and basal DA release SB 243213 : 5-HT2C antagonist Striatum NAC -30 30 60 90 120 150 100 140 160 veh. SB 243213 (1) SB 243213 (3) SB 243213 (10) -30 30 60 90 120 150 100 140 160 veh. SB 243213 (1) SB 243213 (3) SB 243213 (10) *** * ** *** DA (% of baseline) *p<0.05, **p<0.01, ***p<0.001 PLSD test Time (min) Tonic inhibitory control on DA release in both the striatum and NAC

5-HT2C receptors and basal DA release Ro 60-0175: 5-HT2C agonist 60 80 100 120 vehicle 0.3 1 3 mg/kg * ** DA (% of baseline) -30 30 90 150 180 140 NAC striatum Time (min) Ro60-0175 (3 mg/kg) Phasic inhibitory control on DA release in both the striatum and NAC

5-HT2C receptors and basal DA release Conclusions NAC Frontal Cortex Striatum DA release 5-HT2C receptors inhibitory control tonic and phasic SNc VTA Similar in both the striatum and the NAC

Constitutive activity of 5-HT2C receptor and basal DA release 5-HT2C 5-HT DR tonic inhibitory control 5-HT TM1 TM2 TM3 TM4 TM5 TM6 TM7 DA « Activation of 5-HT2C receptors inhibits … DA release in the NAc in a manner which is independent of the inhibition of 5-HT neurons » Willins and Meltzer, Brain Res, 1998 endogenous 5-HT tonic inhibitory control ?

5-HT2C receptors and basal DA release NAC Striatum DA 5-HT2CR antagonists tonic inhibitory control magnitude of effect: SB 206553 >> SB 242084, SB 243213 in vitro studies: SB 206553 SB 242084 SB 243213 different pharmacological properties SNc VTA

Effect of 5-HT2C antagonists on IP accumulation in CHO cells expressing the 5-HT2C receptor 25 50 75 100 125 150 150 150 SB 242084 SB 206553 125 125 SB 206553 + SB 242084 (100 nM) 100 100 IP Accumulation 75 75 (% basal) 50 50 25 25 10 - 11 9 8 7 6 5 10 10 - - 11 11 10 10 - - 10 10 10 10 - - 9 9 10 10 - - 8 8 10 10 - - 7 7 10 10 - - 6 6 10 10 - - 5 5 -log [DRUG] M SB 206553: 5-HT2C inverse agonist In vitro: SB 242084: 5-HT2C weak efficacy agonist

Effect of 5-HT2C antagonists on IP accumulation in CHO cells expressing the 5-HT2C receptor -100 -80 -60 -40 -20 20 IP Accumulation % basal activity 300 nM 1 uM SB 206553 clozapine 30 nM SB 243213 SB 206553: 5-HT2C inverse agonist In vitro: SB 243213: 5-HT2C antagonist

inverse agonist in vivo ? SB 206553 inverse agonist in vivo ? DA release agonist and inverse agonist: - opposite effects - both blocked by antagonists

SB 242084 vs Ro60-0175 SB 242084 (1 mg/kg, ip.) 30 min before Ro 60-0175 (3 mg/kg, ip.) Striatum NAC * * DA ( % of baseline ) ** *** Vehicle + Ro60-0175 SB242084 * p<0.05, ** p<0.01, ***p<0.001 PLSD test SB 242084 inhibits the effect of the 5-HT2CR agonist Ro 60-0175

SB 243213 vs Ro60-0175 SB 243213 (1 mg/kg, ip.) 30 min before Ro 60-0175 (3 mg/kg, ip.) Striatum NAC ** * DA ( % baseline ) *** *** Vehicle + Vehicle + Ro60-0175 SB243213 + Vehicle SB243213 + Ro60-0175 Vehicle + Vehicle + Ro60-0175 SB243213 + Vehicle SB2432123 + Ro60-0175 * p<0.05, ** p<0.01, ***p<0.001 PLSD test SB 243213 inhibits the effect of the 5-HT2CR agonist Ro 60-0175

SB 242084 (1 mg/kg, ip.) 30 min before SB 206553 (5 mg/kg, ip.) SB 242084 vs SB 206553 SB 242084 (1 mg/kg, ip.) 30 min before SB 206553 (5 mg/kg, ip.) Vehicle + SB206553 SB242084 p<0.05, *** p<0.001 vs controls; + p<0.05 vs « SB 206553 » *** * + DA ( % of baseline ) Striatum NAC SB 242084 inhibits the excitatory effect of SB 206553 SB 206553 acts as a 5-HT2C inverse agonist in vivo

SB 243213 vs SB 206553 SB 243213 (1 mg/kg, ip.) 60 min before SB 206553 (5 mg/kg, ip.) DA ( % baseline ) Vehicle + SB206553 SB243213 *** ** Striatum NAC 100 125 150 ** p<0.01, ***p<0.001 PLSD test SB 243213 inhibits the excitatory effect of SB 206553 SB 206553 acts as a 5-HT2C inverse agonist in vivo

SB 206553-induced DA release is unaffected by Striatum NAC SB 206553-induced DA release is unaffected by the reduction of 5-HT extracellular levels by 8-OH-DPAT 8-OH-DPAT (0.1 mg/kg, sc.) 5 min before SB 206553 (5 mgkg, ip.) The effect of SB 206553 occurs independently of endogenous 5-HT

SB 206553-induced DA release is unaffected by the lesion of 5-HT neurons SB 206553 (5 mg/kg, ip.) Striatum Time (min) -30 30 60 90 120 150 DA (% of baseline) 100 125 175 Sham-lesioned 5,7-DHT NAC 175 150 125 100 -30 30 60 90 120 150 The effect of SB 206553 occurs independently of endogenous 5-HT

Constitutive activity of 5-HT2C receptor and basal DA release Conclusions 5-HT2C 5-HT DR tonic inhibitory control 5-HT TM1 TM2 TM3 TM4 TM5 TM6 TM7 DA Constitutive activity of 5-HT2C receptors participates in the tonic inhibitory control of nigrostriatal and mesoaccumbens DA neuron activity in vivo SB 206553: 5-HT2CR inverse agonist in vivo

II. 5-HT2C receptors and antipsychotic drugs constitutive activity basal stimulated ( - ) DA release ? constitutive activity haloperidol clozapine

Haloperidol 0.01 mg/kg Striatum *** *** ** * SB 206553 (5 mg/kg, ip.) veh. 250 SB 206553 225 haloperidol SB 206553 + + + 200 haloperidol DA (% of baseline) 175 *** 150 ** 125 100 -30 30 60 90 120 Time (min) *p<0.05, **p<0.01, ***p<0.001 vs controls; +++p<0.001 vs haloperidol group PLSD test Involvement of constitutive activity of 5-HT2C receptors inverse agonist ≠ antagonist

Involvement of constitutive activity of 5-HT2C receptors Haloperidol 0.01 mg/kg N. Accumbens SB 206553 (5 mg/kg, ip.) SB 242084 (1 mg/kg, ip.) veh. SB 206553 haloperidol veh. SB 242084 haloperidol + + + DA (% of baseline) *** ** ** *** Time (min) **p<0.01, ***p<0.001 vs controls; +++p<0.001 vs haloperidol group PLSD test Involvement of constitutive activity of 5-HT2C receptors importance of inverse agonism in vivo

Haloperidol and SB 243213 Striatum NAC *** *** Haloperidol (0.01 mg/kg, sc.) 30 minutes after SB 243213 (1 mg/kg, ip.) Striatum NAC -60 -30 30 60 90 120 100 125 150 175 200 225 250 veh. SB 243213 haloperidol DA (% of baseline) *** -60 -30 30 60 90 120 100 125 150 175 200 225 250 *** Time (min) ***p<0.001 PLSD test blockade of 5-HT2C receptors : no effect on haloperidol-induced DA release Δ haloperidol effect : related to the constitutive activity of 5-HT2C receptors

5-HT2C receptors and antipsychotic drugs Clozapine nM range inverse agonist in vitro 5-HT2C in vivo ?

Clozapine and DA release in vivo Striatum NAC *** *** *** * *** * *p<0.05, ***p<0.001 PLSD test Clozapine increases DA release in both the striatum and NAC

Clozapine and 5-HT2C receptors Ro 60-0175 (3 mg/kg, ip.) 15 min before clozapine (1 mg/kg, sc.) Striatum NAC *** ** DA ( % of baseline ) *** *** Vehicle + Ro60-0175 + vehicle vehicle + CLOZ Ro60-0175 + CLOZ Vehicle + Ro60-0175 + vehicle vehicle + CLOZ Ro60-0175 + CLOZ **p<0.01, ***p<0.001 PLSD test Clozapine occupies 5-HT2C receptors in vivo

Clozapine: 5-HT2C inverse agonist in vivo ? Ro 60-0175 (3) Striatum 175 SIGNIFICANT INTERACTION CLOZAPINE (1 mg/kg) 150 ** SB Clo V SB / Clo NON SIGNIFICANT INTERACTION Striatum / NAC SB 206553 (5) *** ** 125 SB 242084 SIGNIFICANT INTERACTION ? 100 *** 75 Ro Clo V Ro / Clo 5-HT2C antagonist or inverse agonist properties ? 5-HT2C inverse agonist? Clozapine does not act as a 5-HT2C antagonist

Striatum NAC Clozapine: 5-HT2C inverse agonist in vivo ? SB 242084 (0.3 mg/kg, ip.) 15 min before clozapine (1 mg/kg, sc.) Striatum NAC -30 30 60 90 120 100 140 160 *** ++ veh. SB 242084 clozapine + clozapine -30 30 60 90 120 100 140 160 + *** DA ( % of baseline ) Time (min) ***p<0.001 vs controls; +p<0.05, ++p<0.01 vs clozapine group PLSD test SB 242084 inhibits the effect of clozapine

inverse agonist property in vivo Clozapine DAergic effects (low doses) 5-HT2C NAC Striatum SNc DA DA ATV inverse agonist property in vivo

III. VTA and NAc 5-HT2C receptors DA release DA 5-HT2C Relative contribution of VTA and NAc 5-HT2C receptors in the control of NAc DA release VTA DA 5-HT2C Phasic inhibitory control: 5-HT2C R agonist Ro 60-0175 Tonic inhibitory control: 5-HT2C R inverse agonist SB 206553

VTA and NAc 5-HT2C receptors experimental procedure DA NAc microdialysis probe shell DA reverse dialysis (0.1- 1 µM) SB 242084 5-HT2C 5-HT2C antagonists Injection canula VTA 1 - Ro 60-0175 : 3 mg/kg, ip. SB 242084 (0.1- 0.5 µg / 0.2 µl) SB 243213 DA 2 - SB 206553 : 5 mg/kg, ip. 5-HT2C

SB 242084 1 - Ro 60-0175-induced DA release Intra-VTA injection of 5-HT2C antagonists SB 242084 Time ( min ) SB 242084 prevents the inhibitory effect of Ro 60-0175 ***p<0.001 vs veh, ++p<0.01, +++p<0.001 vs Ro 60-0175 , PLSD test 0.5µg/0.2µl -30 30 60 90 120 80 100 +++ *** veh / veh SB 242084 / veh veh / Ro 60-0175 SB 242084 / Ro 60-0175 NAc DA (% of baseline) 0.1µg/0.2µl ++

SB 243213 1 - Ro 60-0175-induced DA release Intra-VTA injection of 5-HT2C antagonists SB 243213 veh / veh SB 243213 / veh veh / Ro 60-0175 SB 243213 / Ro 60-0175 -60 -30 30 60 90 120 *** +++ NAc DA (% of baseline) 80 100 0.1µg/0.2µl -60 -30 30 60 90 120 +++ *** 80 100 0.5µg/0.2µl ***p<0.001 vs controls; +++p<0.001 vs RO 60-0175, PLSD test Time ( min ) SB 243213 prevents the inhibitory effect of Ro 60-0175 VTA 5-HT2C Rs participate in the phasic inhibitory control of DA release

SB 242084 2 - SB 206553-induced DA release Intra-VTA injection of 5-HT2C antagonists SB 242084 NAc DA (% of baseline) time (min) -30 30 60 90 120 80 100 140 160 180 0.1 µg/0,2µl *** v eh / veh SB 242084 / veh veh / SB 206553 SB 242084 / SB 206553 ***p<0.001 vs veh PLSD test 0.5 µg/0,2µl SB 242084 has no influence on SB 206553-induced DA release VTA 5-HT2C Rs do not participate in the tonic inhibition of DA release related to the CA of 5-HT2C Rs

VTA and NAc 5-HT2C receptors DA DA release 5-HT2C Nucleus Accumbens 5-HT2C receptors VTA DA (-) 5-HT2C

SB 242084 1 - Ro 60-0175-induced DA release Intra-NAc perfusion of 5-HT2C antagonist SB 242084 veh / veh SB 242084 / veh veh / Ro 60-0175 SB 242084 / Ro 60-0175 NAc DA (% of baseline) -30 30 60 90 120 80 100 ** ++ 0.1 µM -30 30 60 90 120 80 100 *** +++ 1 µM Time (min) **p<0.01, ***p<0.001 vs veh; ++p<0.01; +++p<0.001 vs RO 60-0175 group PLSD test SB 242084 prevents the inhibitory effect of Ro 60-0175 NAc 5-HT2C Rs participate in the phasic inhibitory control of DA release

SB 242084 2 - SB 206553-induced DA release Intra-NAc perfusion of 5-HT2C antagonists SB 242084 v eh / veh SB 242084 / veh veh / SB 206553 SB 242084 / SB 206553 -30 30 60 90 120 100 140 160 *** +++ -30 30 60 90 120 100 140 160 *** +++ NAc DA (% of baseline) 0.1µM 1µM time (min) ***p<0.001 vs veh; +++p<0.001 vs SB 206553 PLSD test SB 242084 prevents the excitatory effect of SB 206553 NAc 5-HT2C Rs participate in the tonic inhibitory control exerted by the CA of 5-HT2C Rs

SB 206553 Basal DA release intra-VTA and intra-NAc administration -30 30 60 90 120 100 time (min) 0.5 µg/0.2 µl 110 130 time (min) -30 30 60 90 120 100 ** 1µM 110 130 veh SB 206553 NAc DA (% of baseline) **p<0.01 vs veh test PLSD Intra-NAc application of the 5-HT2C inverse agonist SB 206553 increases basal DA release in the NAc

VTA and NAc 5-HT2C receptors DA DA release 5-HT2C phasic & tonic VTA and NAc 5-HT2C Rs both participate in the overall inhibitory control of NAc DA release The NAc may represent a primary site of action for the effects of the CA of 5-HT2C Rs VTA ≠ levels of 5-HT2C R CA in the NAc and VTA related to RNA editing DA lowers the CA and modulates the desensitization state of 5-HT2C Rs generates tissue-specific expression of 5-HT2C R isoforms with distinct biological properties (Niswender et al., 1999; Marion et al., 2004) 5-HT2C phasic

5-HT2C receptor control of DA ascending pathways in vivo Conclusions 5-HT2C receptor DA “fine tuning” DA pathways NAC / Striatum inhibitory tonic / phasic control involvement of constitutive activity in vivo - DAergic effects of antipsychotic drugs - clozapine: inverse agonist in vivo DA region-dependent control by constitutive activity - NAc → primary site of action VTA SN - Role of mRNA editing ( ↓ constitutively active isoforms) in the control of DA neuron excitability

Conclusions DA 5-HT2C receptor 5-HT2C receptor control of DA ascending pathways in vivo Conclusions 5-HT2C receptor DA “fine tuning” DA pathways NAC / Striatum need for studies assessing the functional significance and the therapeutic potential of inverse agonism in vivo DA VTA SN Long term studies …… studies in animal models of Parkinson’s disease….

« Neuropharmacologie et Neurochimie Fonctionnelle » NAc Université Bordeaux 2 INSERM U 862 France DA 5-HT2C Umberto Spampinato Philippe De Deurwaerdère Guillaume Lucas Grégory Porras Sylvia Navailles Delphine Moison Dimitri Ryczko VTA DA Pr William Clarke & Dr Kelly Berg 5-HT2C Department of Pharmacology, University of Texas Health Science Center, San Antonio, USA

    DA DA SNc VTA 5-HT2A 5-HT3 5-HT4 5-HT receptors and in vivo DA release: conditional involvement DA n. Accumbens VTA 5-HT2C  DA cell firing Striatum SNc DA  5-HT2A  DA synthesis  DA transmission   5-HT tone  DA cell firing 5-HT3   DA transmission  DA cell firing 5-HT4 Factors ? state-dependent facilitatory control tonic inhibitory control Selective modulation of one DA pathway depending on the state of DA neuron activity Rational basis for therapeutics

5-HT and Parkinson’s disease Preclinical data (rat): central 5-HT transmission 5-HT2C antagonism Catalepsy central 5-HT transmission (SSRI) 5-HT4 agonists Catalepsy 5-HT2C antagonism motor response (L-dopa, DA agonists) Clinical data: 5-HT4 agonist (cisapride) Park (tremor) central 5-HT transmission (SSRI) Park symptoms dyskinesia 5-HT1A agonism (buspiron) dyskinesia 5-HT2A/2C - 5-HT3 antagonists Dopa-induced psychosis (no change motor score) (mianserin, ondansetron) atypical antipsychotic (clozapine) Dopa-induced psychosis, tremor 5-HT2/3 + α2 antagonism (mirtazapine) tremor- dyskinesia

DA release & cocaine-induced effects 5-HT2C receptors: DA release & cocaine-induced effects control / contribution involvement VTA 5-HT2C R NAc 5-HT2C R yes Inhibitory effect on DA release yes yes Phasic inhibitory control of DA release yes no Basal DA release no yes (ago) Cocaine-induced DA release yes (ago/antag) yes (ago) Cocaine-induced behaviors no (ago/antag) Involvement in the systemic effect of 5-HT2C agents no DA release yes (antag) no Behaviors yes (antag)

DA release & cocaine-induced behaviors 5-HT2C receptors: DA release & cocaine-induced behaviors ip. Intra-VTA Intra-NAc 5-HT2C agents administration agonist antagonist basal behaviors agonist antagonist Basal DA release Ro-induced DA antagonist agonist antagonist Cocaine- DA agonist antagonist Cocaine- behaviors

5-HT2C receptors and DA neuron firing rate SB 242084 Ro 60-0175 VTA *p<0.05, **p<0.01 vs veh, Tukey’s test Di Matteo et al., 1999 Tonic and phasic inhibitory control on DA neuron activity

Conclusions (+/-) (-) DA transmission release DA NAc Change of 5-HT2C Change of behavior cellular activity 5-HT2C receptor control of the mesoaccumbens DA pathway: VTA composite responses involving different populations of 5-HT2C receptors in multiple brain nuclei DA controlling NAc DA release 5-HT2C and/or DA transmission

5-HT2C Rs control DA release and/or transmission biochemical electrophysiological behavioral studies Fr Cortex / NAc DA transmission DA release 5-HT2C “DARP-32” Svenningsson et al., Pnas, 2001 regulation of DA exocytosis via DA neuron firing 5-HT2C VTA DA firing 5-HT2C modulation of DA transmission may occur independently from changes of DA release

5-HT2C receptors and basal DA release SB 206553 - SB 242084 Frontal Cortex Gobert et al., 2000 TIME (MIN) PERCENT OF CONTROL * p<0.05 ANOVA SB 206553 SB 242084

5-HT2C receptors and basal DA release Ro 60-0175 Frontal Cortex TIME (MIN) PERCENT OF CONTROL Gobert et al., 2000 * p<0.05 ANOVA

Striatum NAC Clozapine: 5-HT2C inverse agonist in vivo ? SB 243213 (1 mg/kg, ip.) 30 min before clozapine (1 mg/kg, sc.) Striatum NAC -60 -30 30 60 90 120 100 140 160 *** ** veh. SB 243213 clozapine + clozapine -60 -30 30 60 90 120 100 140 160 *** DA ( % of baseline ) Time (min) **p<0.01, ***p<0.001 vs controls PLSD test SB 243213 inhibits the effect of clozapine