Cloning and fnctional expression of an apparent splice variant of the murine 5-HT3 receptor A subunit

Hope, Anthony G.; Downie, David L.; Sutherland, Lesley; Lambert, Jeremy J.; Peters, John A.; Burchell, Brian

doi:10.1016/0922-4106(93)90128-v

Cited by 152 publications

(108 citation statements)

References 8 publications

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“…In particular, slow rates of desensitization were apparent in nicotine-induced increases in presynaptic Ca2l evel measured at synapses between spinal cord neurons and lumbar sympathetic ganglia (Scheggenburger et al, 1993). Slow desensitization rates were also observed for 5-HT 3 receptor-induced responses in differentiated neuroblastoma x glioma NG1O8-15 cells (Shao et al, 1991;Hope et al, 1993), which display many of the features of PNS cholinergic neurons, including neuritic processes having numerous synapticlike varicosities (Nelson et al, 1976), as compared with undifferentiated cells. Such slow kinetics would indicate a function for 5-HT3 receptors at presynaptic sites that is likely quite distinct from that observed postsynaptically for these receptors.…”

Section: Discussionmentioning

confidence: 90%

High Calcium Permeability of Serotonin 5‐HT₃ Receptors on Presynaptic Nerve Terminals from Rat Striatum

Rondé

Nichols

1998

Journal of Neurochemistry

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Abstract:The serotonin 5-HT3 receptor, a ligand-gated ion channel, has previously been shown to be present on a subpopulation of brain nerve terminals, where, on activation, the 5-HT3 receptors induce Ca 2~influx. Whereas postsynaptic 5-HT 3 receptors induce depolarization, being permeant to Naãnd K~, the basis of presynaptic 5-HT3 receptor-induced calcium influx is unknown. Because the small size of isolated brain nerve terminals (synaptosomes) precludes electrophysiological measurements, confocal microscopic imaging has been used to detect calcium influx into them. Application of 100 n M 1-(m-chlorophenyl) biguanide (mCPBG), a highly specific 5-HT3 receptor agonist, induced increases in internal free Ca 2~concentration ([Ca2~]) and exocytosis in a subset of corpus striatal synaptosomes. mCPBG-induced increases in [Ca2~]ranged from 1.3 to 1 .6 times over basal values and were inhibited by 10 nM tropisetron, a potent and highly specific 5-HT 3 receptor antagonist, but were insensitive to the removal of external free Na~(substituted with N-methyl-o-glucamine), to prior depolarization induced on addition of 20 mM K~, or to voltage-gated Ca 2~channel blockade by 10 p~M Co2~/Cd2õrbyl ,uMw-conotoxin MVIIC/1~tMw-conotoxin GVIA/200 nM agatoxin TK. In contrast, the Ca2ĩ nflux induced by 5-HT 3 receptor activation in NG1 08-15 cells by 1 ,tiM mCPBG was substantially reduced by 10.tMCo 2~/Cd2ãnd was completely blocked by 1 ,uM nitrendipine, an L-type Ca2~channel blocker. We conclude that in contrast to the perikaryal 5-HT 3 receptors, presynaptic 5-HT3 receptors appear to be uniquely calcium-permeant. Key Words: Serotonin 5-HT3 receptorPresynaptic regulation-Synaptosomes-Confocal microscopy-Internal free Ca 2~concentration measurements-NG1O8-15 cells.

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Section: Discussionmentioning

confidence: 90%

High Calcium Permeability of Serotonin 5‐HT₃ Receptors on Presynaptic Nerve Terminals from Rat Striatum

Rondé

Nichols

1998

Journal of Neurochemistry

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“…To study the effects of 5-HI, we examined whole-cell currents activated by 5-HT in N1E-115 neuroblastoma cells expressing the native mouse 5-HT 3 receptor (Neijt et al, 1988;Hope et al, 1993), and subsequently in HEK 293 cells transiently expressing the recombinant mouse 5-HT 3A receptor. Inward current with a rapid onset was observed upon application of a maximally efficacious concentration of 5-HT (30 μM) for 10 s to either N1E-115 cells or transfected HEK 293 cells (Fig.…”

Section: Positive Modulation Of 5-ht 3 Receptor-mediated Current By 5-himentioning

confidence: 99%

The L293 residue in transmembrane domain 2 of the 5-HT3A receptor is a molecular determinant of allosteric modulation by 5-hydroxyindole

Lovinger

2008

Neuropharmacology

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Allosteric modulation of ligand-gated ion channels can play important roles in shaping synaptic transmission. The function of the 5-hydroxytryptamine (serotonin) type 3 (5-HT 3 ) receptor, a member of the Cys-loop ligand-gated ion channel superfamily, is modulated by a variety of compounds such as alcohols, anesthetics and 5-hydroxyindole (5-HI). In the present study, the molecular determinants of allosteric modulation by 5-HI were explored in N1E-115 neuroblastoma cells expressing the native 5-HT 3 receptor and HEK 293 cells transfected with the recombinant 5-HT 3A receptor using molecular biology and whole-cell patch-clamp techniques. 5-HI potentiated 5-HT-activated currents in both N1E-115 cells and HEK 293 cells, and significantly decreased current desensitization and deactivation. Substitution of Leu293 (L293, L15′) in the second transmembrane domain (TM2) with cysteine (L293C) or serine (L293S) abolished 5-HI modulation. Other mutations in the TM2 domain, such as D298A and T284F, failed to alter 5-HI modulation. The L293S mutation enhanced dopamine efficacy and converted 5-HI into a partial agonist at the mutant receptor. These data suggest that 5-HI stabilizes the 5-HT 3A receptor in the open state by decreasing both desensitization and 5-HT unbinding/channel closing; and L293 is a common site for both channel gating and allosteric modulation by 5-HI. Our observations also indicate existence of a second 5-HI recognition site on the 5-HT 3A receptor which may overlap with the 5-HT binding site and is not involved in the positive modulation by 5-HI. These findings support the idea that there are two discrete sites for 5-HI allosteric modulation and direct activation in the 5-HT 3A receptor.

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“…HEK-5-HT3-As cells (Hope et al, 1993;Downie et al, 1995) were cultured in minimal essential medium containing 10% v/v foetal calf serum, supplemented with streptomycin (40 jig m['), penicillin (40 units (0.75 mg ml-') and harvested between passage 3 and 40. Pelleted cells were frozen at -80°C prior to assay (less than 2 months).…”

Section: Cell Culturementioning

confidence: 99%

Evidence that the atypical 5‐HT₃ receptor ligand, [³H]‐BRL46470, labels additional 5‐HT₃ binding sites compared to [³H]‐granisetron

Steward

Jin

Bentley

et al. 1995

British J Pharmacology

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Cloning and fnctional expression of an apparent splice variant of the murine 5-HT3 receptor A subunit

Cited by 152 publications

References 8 publications

High Calcium Permeability of Serotonin 5‐HT₃ Receptors on Presynaptic Nerve Terminals from Rat Striatum

High Calcium Permeability of Serotonin 5‐HT₃ Receptors on Presynaptic Nerve Terminals from Rat Striatum

The L293 residue in transmembrane domain 2 of the 5-HT3A receptor is a molecular determinant of allosteric modulation by 5-hydroxyindole

Evidence that the atypical 5‐HT₃ receptor ligand, [³H]‐BRL46470, labels additional 5‐HT₃ binding sites compared to [³H]‐granisetron

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Cloning and fnctional expression of an apparent splice variant of the murine 5-HT3 receptor A subunit

Cited by 152 publications

References 8 publications

High Calcium Permeability of Serotonin 5‐HT3 Receptors on Presynaptic Nerve Terminals from Rat Striatum

High Calcium Permeability of Serotonin 5‐HT3 Receptors on Presynaptic Nerve Terminals from Rat Striatum

The L293 residue in transmembrane domain 2 of the 5-HT3A receptor is a molecular determinant of allosteric modulation by 5-hydroxyindole

Evidence that the atypical 5‐HT3 receptor ligand, [3H]‐BRL46470, labels additional 5‐HT3 binding sites compared to [3H]‐granisetron

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High Calcium Permeability of Serotonin 5‐HT₃ Receptors on Presynaptic Nerve Terminals from Rat Striatum

High Calcium Permeability of Serotonin 5‐HT₃ Receptors on Presynaptic Nerve Terminals from Rat Striatum

Evidence that the atypical 5‐HT₃ receptor ligand, [³H]‐BRL46470, labels additional 5‐HT₃ binding sites compared to [³H]‐granisetron