Actions of 5-hydroxytryptamine and 5-HT1A receptor ligands on rat dorso-lateral septal neurones in vitro.

1. The actions of 5-hydroxytryptamine (5-HT) and some 5-HT1A receptor ligands on neurones in the rat dorso-lateral septal nucleus were recorded in vitro by intracellular recording techniques. 2. In the presence of tetrodotoxin (1 microM) to block any indirect effects, bath application of 5-HT (0.3-30 microM) hyperpolarized the neurones in a concentration-dependent manner and reduced membrane resistance. The hyperpolarization did not exhibit desensitization and was sometimes followed by a small depolarization. 3. The 5-HT1A receptor ligands, 8-hydroxy-2-(di-n-propylamino) tetralin (8-OH-DPAT), N,N-dipropyl-5-carboxamidotryptamine (DP-5-CT) and buspirone but not the non-selective 5-HT1 receptor agonist, 1-m-trifluoromethylphenylpiperazine (TFMPP), also hyperpolarized the neurones. 4. 5-HT, 8-OH-DPAT and DP-5-CT appeared to act as full agonists whereas buspirone behaved as a partial agonist. The estimated EC50S were: DP-5-CT 15 nM, 8-OH-DPAT 110 nM, 5-HT 3 microM and buspirone 110 nM. 5. At a concentration of 3 microM, the putative 5-HT1A receptor antagonists, spiperone, methiothepin, NAN-190 (1-(2-methoxyphenyl)-4-[4-(2-pthalimido)butyl]piperazine) and MDL 73005EF (8-[2-(2,3-dihydro-1,4-benzodioxin-2-yl-methylamino)ethyl]-8- azaspiro[4,5]decane-7,9-dione methyl sulphonate), produced a parallel rightward shift in the concentration-response curve to 5-HT with no significant reduction in the maximum response. The estimated pA2 values were: NAN-190 6.79, MDL 73005EF 6.59, spiperone 6.54 and methiothepin 6.17.(ABSTRACT TRUNCATED AT 250 WORDS)     

Centrally acting hypotensive agents with affinity for 5-HT1A binding sites inhibit forskolin-stimulated adenylate cyclase activity in calf hippocampus.

1. A number of centrally acting hypotensive agents and other ligands with high affinity for 5-hydroxytryptamine1A (5-HT1A) recognition sites have been tested on forskolin-stimulated adenylate cyclase activity in calf hippocampus, a functional model for 5-HT1A-receptors. 2. Concentration-dependent inhibition of forskolin-stimulated adenylate cyclase activity was elicited by the reference 5-HT1-receptor agonists (mean EC50 value, nM): 5-HT (22), 5-carboxamidotryptamine (5-CT, 3.2), 8-hydroxy-2-(di-n-propylamino)-tetralin (8-OH-DPAT, 8.6), N,N-dipropyl-5-carboxamidotryptamine (DP-5-CT, 2.3), 1-[2-(4-aminophenyl)ethyl]-4-(3-trifluoromethylphenyl)-piperazine (PAPP or LY 165163, 20), 5-methoxy-3-(1,2,3,6-tetrahydro-4-pyridinyl)-1H indole (RU 24969, 20), buspirone (65) and ipsapirone (56). Emax amounted to 18-20% inhibition for all but the latter two agonists (14%). 3. The following hypotensive agents with high affinity for 5-HT1A sites were potent agonists in this system (mean EC50 value, nM): flesinoxan (24), indorenate (99), erythro-1-(1-[2-(1,4-benzodioxan-2-yl)-2-hydroxyethyl]-4-piperidyl )- 2-benzimidazolinone (R 28935, 2.5), urapidil (390) and 5-methyl-urapidil (3.5). The first two agents were full agonists, whereas the latter three acted as partial agonists with 60-80% efficacy. 4. Metergoline and methysergide behaved as full agonists and cyanopindolol as a partial agonist with low efficacy. Spiroxatrine and 2-(2,6-dimethoxyphenoxyethyl)aminomethyl- 1,4-benzodioxane (WB 4101) which bind to 5-HT1A sites with nanomolar affinity, were agonists and inhibited potently forskolin-stimulated adenylate cyclase in calf hippocampus, showing mean EC50 values of 23 and 15 nM, respectively. Spiroxatrine and WB 4101 yielded 90% and 50% efficacy, respectively. 5. Spiperone and methiothepin (each 1 microM) caused rightward shifts of the concentration-effect curve to 8-OH-DPAT, without loss of the maximal effect, as did the partial agonist cyanopindolol (0.1 microM) and the (-)- and (+)-enantiomers of pindolol (1 microM and 0.1 mM, respectively). 6. There was an excellent correlation (r = 0.90, P = 0.0001) between the pEC50 values (ranging from 6.4 to 8.7) of the 19 agonists tested at adenylate cyclase and their pKD for 5-HT1A recognition sites. Apparent pKB values of antagonists at adenylate cyclase and their pKD values for 5-HT1A binding sites were also significantly correlated. 7. This study further indicates that the 5-HT1A recognition site and the 5-HT receptor mediating inhibition of adenylate cyclase in hippocampus are the same.(ABSTRACT TRUNCATED AT 400 WORDS)     

Central 5-HT1A receptors and the mechanism of the central hypotensive effect of (+)8-OH-DPAT, DP-5-CT, R28935, and urapidil

This study investigated the central hypotensive effects of drugs that possess a high affinity for central 5-hydroxytryptamine (5-HT1A) binding sites; (+)8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT), N,N-dipropylcarboxamidotryptamine (DP-5-CT), erythro-1-(1-[2(1,4-benzodioxan-2-yl)-2-hydroxyethyl]- 4-piperidyl)-2-benzimidazolinone (R28935) and urapidil proved to possess high affinity and selectivity for central 5-HT1A binding sites, labeled by [3H]8-OH-DPAT. (+)8-OH-DPAT (0.1-10 micrograms/kg) given through the left vertebral artery of chloralose-anesthetized cats, lowered blood pressure by a biphasic dose-response curve. When given systemically, 10- to 100-fold higher doses of (+)8-OH-DPAT were necessary to obtain the same hypotensive effect when compared with central administration. Besides 8-OH-DPAT, R28935, DP-5-CT and urapidil also lowered blood pressure by a central mechanism in doses that were ineffective when given systemically. The central hypotensive effect of 0.3 micrograms/kg (+)8-OH-DPAT, 3 micrograms/kg DP-5 CT, and 3 micrograms/kg R28935 could be blocked by 100 micrograms/kg (-)pindolol, indicating that central 5-HT1A receptors are involved. High doses of (+)8-OH-DPAT (3-10 micrograms/kg) can also lower blood pressure by activating central alpha 2-adrenoceptors. The hypotensive effect of 300 micrograms/kg urapidil given through the vertebral artery could not be blocked by (-)pindolol. These results indicate the involvement of central 5-HT1A receptors in the mechanism of the central hypotensive activity of (+)8-OH-DPAT, DP-5-CT, and R28935.     

5-HT1A-receptors mediate stimulation of adenylate cyclase in rat hippocampus

Serotonin (5-HT) stimulated adenylate cyclase activity in homogenates of rat hippocampus. This effect was pharmacologically characterised with a series of agonists and antagonists of various structural classes. These compounds where also tested in radioligand binding studies using selective ligands for the various subtypes of 5-HT1 and 5-HT2 receptors. 5-HT1A, 5-HT1B and 5-HT1C recognition sites were labelled with [3H]8-OH-DPAT([3H]8-hydroxy-2-(di-n-propylamino)-tetralin) in pig cortex membranes, [125I]CYP([125I]iodocyanopindolol) in rat cortex and [3H]mesulergine in pig choroid plexus membranes, respectively. The rank order of potency of 13 agonists stimulating adenylate cyclase activity in homogenates of rat hippocampus was in good agreement with the rank order of affinity of these agonists for the 5-HT1A binding site: N,N-dipropyl-5-carboxamidotryptamine (DP-5-CT) greater than 5-carboxamidotryptamine (5-CT) greater than 8-OH-DPAT greater than 5-HT greater than 5-methoxytryptamine (5-OCH3T) greater than d-LSD greater than 5-methoxy-3-(1,2,3,6-tetrahydro-4-pyridinyl)-1H-indole (RU 24969) greater than alpha-methylserotonin (alpha-CH3-5-HT) greater than dopamine greater than 2-methylserotonin (2-CH3-5-HT). The correlation between the respective potencies and affinities of these agonists was r = 0.934, P less than 0.001. There was no correlation between stimulation of adenylate cyclase activity by these agonists and their affinity for 5-HT1B, 5-HT1C or 5-HT2 binding sites. r = 0.381-0.108, P less than 0.20-0.73.(ABSTRACT TRUNCATED AT 250 WORDS)     

GH4ZD10 cells expressing rat 5-HT1A receptors coupled to adenylyl cyclase are a model for the postsynaptic receptors in the rat hippocampus.

1. Vasoactive intestinal polypeptide (VIP) stimulated adenosine 3':5'-cyclic monophosphate (cyclic AMP) production by cultured GH4ZD10 cells with an EC50 value of about 7 nM. The extracellularly recovered cyclic AMP predominated, and was reduced by co-incubation with 8-hydroxy-2-(di-n-propyl-amino) tetralin (8-OH-DPAT) and 5-hydroxytryptamine (5-HT), whereas dopamine (0.1-30 microM) did not reduce VIP-stimulated cyclic AMP production. 2. The responses to 5-HT and 8-OH-DPAT were blocked by (-)-alprenolol and NAN 190. The antagonism by (-)-alprenolol was competitive in nature with a pA2 value of 7.0. 3. The responsiveness of the cells to 5-HT agonists was highly dependent upon the culturing conditions used. Thus, 8-OH-DPAT inhibition of VIP (30 nM)-stimulated cyclic AMP production decreased with increasing passage number of the cells. Reduction of the zinc concentration used to promote expression of the 5-HT1A receptor gene produced a greater sensitivity of the cells to 5-HT agonists. 4. Under such conditions, the following efficacies (5-HT = 100) were found: lisuride 106, (+)-lysergic-acid diethylamide 100, 5-methoxy-N,N-dimethyltryptamine 98, RU 24949 98, 5-carboxamidotryptamine 97, (+/-)-8-OH-DPAT 90, (+)-8-OH-DPAT 87, 1-[2-(4-aminophenyl)ethyl]-4-(3-trifluoromethylphenyl)-piperazine 86, flesinoxan 79/88, (-)-8-OH-DPAT 62, buspirone 43/50, ipsapirone 46. Spiroxatrine and spiperone had a low intrinsic activity, but reduced the response to 5-HT. These efficacies are similar to those reported in the literature for post-synaptically localized 5-HT1A receptors in the rat hippocampus.(ABSTRACT TRUNCATED AT 250 WORDS)     

Vasodilatation elicited by 5-HT1A receptor agonists in constant-pressure-perfused rat kidney is mediated by blockade of alpha 1A-adrenoceptors.

The vasodilator mechanism of the putative serotonin1A (5-HT) receptor agonists, urapidil, 5-methyl-urapidil, ipsapirone, flesinoxan and 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT) was investigated in constant-pressure perfused rat kidneys. The compounds (10(-12)-10(-7) mol bolus injection) neither enhanced basal flow nor evoked vasodilatation in kidneys preconstricted by 27 mM KCl, 1.5 mM BaCl2 or 10(-6) M prostaglandin (PG)F2 alpha, but evoked a dose-dependent, reversible and spiroxatrine-resistant increase in vasodilatation of organs preconstricted by 6 x 10(-7) M noradrenaline. 5-Carboxamidotryptamine and sumatriptan did not reverse the vasoconstriction induced by all stimuli or that induced by noradrenaline in the presence of 5-HT2 plus 5-HT3 receptor blockade. No correlation for the vasorelaxant drugs was found between their -log ED50 in rat kidney and pKi values at 5-HT1A binding sites in pig cortex as determined in radioligand experiments. The relaxation in rat kidney induced by 5-HT1A receptor agonists and alpha 1A-adrenoceptor-selective antagonists (WB 4101 and (+)-niguldipine) was significantly correlated with pKi values at alpha 1A binding sites in rat cortex and the pA2 values derived from contraction studies for competitive antagonism at alpha 1-adrenoceptors in prostatic portions of the rat vas deferens, but differed from pKi values for alpha 1B binding sites in rat cortex. Thus, the vasodilator effect of the 5-HT1A receptor agonists urapidil, 5-methyl-urapidil, ipsapirone, flesinoxan and 8-OH-DPAT in the noradrenaline-perfused rat kidney appears to be mediated by their concomitant alpha 1A-adrenoceptor blockade. No evidence for a vasodilator effect mediated through 5-HT1A receptors was found under our experimental conditions.     

5-Hydroxytryptamine receptor agonists influence calcium-stimulated adenylate cyclase activity in the cerebral cortex and hippocampus of the rat

The effects of 5-hydroxytryptamine (5-HT) receptor agonists on calcium (Ca2+)-stimulated adenylate cyclase activity in the hippocampus and cerebral cortex of the rat were studied. In the presence of Ca2+ (1.5 microM), 5-HT dose dependently inhibited adenylate cyclase activity (EC50 = 10 +/- 2 nM). The inhibitory effect of 5-HT on Ca2(+)-stimulated adenylate cyclase was antagonized by spiperone (KB = 2 +/- 0.8 nM). The rank order of potency of 5-HT agonists to inhibit Ca2(+)-stimulated adenylate cyclase in the hippocampus was: 5-carboxamidotryptamine (5-CT) greater than 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT) greater than 5-hydroxytryptamine (5-HT) = 5-methoxytryptamine (5-OCH3-T) greater than trifluoromethylphenylpiperazine (TFMPP) greater than m-chlorophenylpiperazine (mCPP). 2-Methyl-5-hydroxytryptamine (2-CH3-5-HT) did not exert an effect on Ca2(+)-stimulated enzyme activity. In the cerebral cortex 5-HT exerted a biphasic stimulatory effect on adenylate cyclase activity in the absence of Ca2+ (EC50 = 0.2 +/- 0.04 nM and 10 +/- 3 microM), whereas 8-OH-DPAT, 5-CT and 2-CH3-5-HT exerted a monophasic effect. In the presence of Ca2+ (1.5 microM), low concentrations of 5-HT, 8-OH-DPAT, 5-CT and 2-CH3-5-HT potentiated adenylate cyclase activity, whereas higher concentrations, except 2-CH3-5-HT, inhibited the enzyme activity. We propose that the 5-HT receptor mediating inhibition of Ca2(+)-stimulated adenylate cyclase in the rat hippocampus corresponds to the 5-HT1A subtype.(ABSTRACT TRUNCATED AT 250 WORDS)     

Lack of apparent receptor reserve at postsynaptic 5-hydroxytryptamine1A receptors negatively coupled to adenylyl cyclase activity in rat hippocampal membranes.

Previous studies have demonstrated the existence of a large receptor reserve for agonists at somatodendritic 5-hydroxytryptamine1A (5-HT1A) serotonin receptors in the raphe nuclei of the rat. 5-HT1A agonists with anxiolytic properties (e.g., buspirone, gepirone, and ipsapirone) display full intrinsic activity at these receptors but are partial agonists at postsynaptic 5-HT1A receptors, which suggests that the latter sites may be devoid of a receptor reserve. In the present studies, this was directly determined by examining the relationship between receptor occupancy and response at postsynaptic 5-HT1A receptors, in rat hippocampus, mediating the inhibition of forskolin-stimulated adenylyl cyclase activity, using the method of partial irreversible receptor inactivation. Rats were treated with vehicle or the irreversible antagonist N-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline (EEDQ), and 24 hr later hippocampi were removed for saturation analysis of [3H]8-hydroxy-2-(di-n-propylamino)-tetralin (8-OH-DPAT) binding to 5-HT1A receptors or for adenylyl cyclase assays. EEDQ (1 and 6 mg/kg) dose-dependently reduced the maximal number of [3H]8-OH-DPAT binding sites by 68.5 and 80%, respectively, without altering the Kd. Concentration-response curves were generated for inhibition of forskolin-stimulated adenylyl cyclase activity by 5-HT and the selective 5-HT1A agonist N,N-dipropyl-5-carboxamidotryptamine (DP-5-CT). EEDQ treatment dose-dependently reduced the maximal inhibitory effect of 5-HT [percentage of inhibition: control, 23.6; EEDQ (1 mg/kg), 13.4; EEDQ (6 mg/kg), 8.9], without altering either the slope factor (1.01) or the EC50 (96.4 nM). Analogous results were obtained with DP-5-CT [percentage of maximal inhibition: control, 24.1; EEDQ (1 mg/kg), 15.2; EEDQ (6 mg/kg), 10.7), again without changes in slope factor (0.89) or EC50 (9.9 nM). Analysis of double-reciprocal plots of equieffective concentrations of agonist, followed by calculation of fractional receptor occupancy, revealed a linear relationship between receptor occupancy and response for both 5-HT and DP-5-CT (i.e., an absence of receptor reserve). The receptor specificity of the effect of EEDQ was demonstrated in two ways. First, it was shown that pretreatment of rats with the selective 5-HT1A partial agonist BMY 7378 (10 mg/kg) before EEDQ afforded substantial protection (about 75%) against loss of the inhibitory effect of DP-5-CT on forskolin-stimulated adenylyl cyclase activity. Second, EEDQ did not alter the inhibition of forskolin-stimulated adenylyl cyclase activity induced by the adenosine A1 receptor agonist phenylisopropyladenosine (PIA).(ABSTRACT TRUNCATED AT 400 WORDS)     

Effects of 5-HT and 5-HT1A receptor agonists and antagonists on dorsal vagal preganglionic neurones in anaesthetized rats: an ionophoretic study.

1. Effects of ionophoretic administration of 5-hydroxytryptamine (5-HT) and selective 5-HT1A receptor agonists and antagonists on identified dorsal vagal preganglionic and dorsal raphe neurones were studied in pentobarbitone sodium or chloral hydrate-anaesthetized rats, respectively. 2. Extracellular recordings were made from 176 preganglionic neurones in the dorsal vagal nucleus (DVN). Application of 5-HT at low currents (< or = 10 nA) increased the activity of these neurones. However, at increased currents (10-60 nA), it had a predominantly depressant effect. Application of selective 5-HT1A receptor antagonists, (+/-)-pindolol or WAY-100635, attenuated the excitatory responses evoked by 5-HT. 3. Ionophoresis of the 5-HT1A receptor agonist, 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT) (5-30 nA) increased the firing rate of 19 and decreased that of 67 of the 104 vagal neurones tested. Other 5-HT1A receptor agonists, flesinoxan and N,N-di-n-propyl-5-carboxamidotryptamine (DP-5-CT) also had predominantly depressant effects. 4. (+/-)-Pindolol attenuated excitations but not inhibitions evoked by 8-OH-DPAT. Surprisingly, WAY-100635 and 8-OH-DPAT produced the same effect on these neurones and when applied together, WAY-100635 failed to attenuate the 8-OH-DPAT responses. 5. Dorsal raphe neurones were identified by their low, regular firing rate and their subsequent histological localization. 8-OH-DPAT reversibly reduced the activity in all 7 neurones tested and this was antagonized by WAY-100635 in all 3 neurones tested. 6. In conclusion, 5-HT applied to vagal preganglionic neurones evokes excitatory and inhibitory responses. The excitatory, but not the inhibitory responses may be mediated, at least in part, by activation of 5-HT1A receptors.     

5-CT stimulation of adenylyl cyclase activity in guinea-pig hippocampus: evidence for involvement of 5-HT7 and 5-HT1A receptors.

1. A number of compounds, including the selective 5-HT7 receptor antagonist SB-258719, were investigated for their effect on [3H]-5-carboxamidotryptamine (5-CT) radioligand binding and 5-CT-stimulated adenylyl cyclase activity in guinea-pig hippocampal membranes, in order to confirm the presence of functionally coupled 5-HT7 receptors in this tissue. 2. The [3H]-5-CT radioligand binding profile was consistent with binding predominantly to 5-HT7 receptors. The affinity of SB-258719 (pKi 7.2+/-0.1) was similar to its reported human 5-HT7 receptor affinity. 3. In the adenylyl cyclase functional assay, 5-CT was a potent and full agonist compared to 5-HT, whereas 8-hydroxy-dipropylaminotetralin (8-OH-DPAT) was a partial agonist (intrinsic activity 0.4+/-0.1). The rank order of potency for agonists (5-CT>5-HT approximately 8-OH-DPAT) was consistent with activation of 5-HT7 receptors. SB-258719 (5 microM) and methiothepin (1 microM) surmountably antagonized the response to 5-CT, consistent with competitive antagonism. The pKB for SB-258719 (7.2+/-0.1) was in good agreement with its reported antagonist potency at the human cloned 5-HT7 receptor. 4. In the functional assay, WAY-100635 (100 nM) and cyanopindolol (1 microM) induced a biphasic 5-CT response curve, consistent with selective antagonism of a component of the response to 5-CT. The estimated pKB values for WAY-100635 and cyanopindolol (9.6 and 8.4 respectively) were in good agreement with their reported 5-HT1A receptor affinities. 5. The data are consistent with the presence of 5-HT7 receptors in guinea-pig hippocampus which are positively coupled to adenylyl cyclase. In addition, 5-HT7 receptor-mediated stimulation of adenylyl cyclase activity in this tissue appears to be augmented by a mechanism involving 5-HT1A receptor activation.     

The septum and the hippocampus differentially mediate anxiolytic effects of R(+)-8-OH-DPAT

Infusing the 5-HT1A receptor agonist R(+)-8-OH-DPAT into the septum or hippocampus reduced the fear responses of rats differentially in the elevated plus-maze and shock-probe burying tests, two rat models of anxiety. Intra-septal infusions of R(+)-8-OH-DPAT (5 and 10 microg) produced dramatic reductions in rat burying behavior in the shock-probe test, whereas it did not alter rat open-arm activity in the plus-maze test, across a wide range of doses (0.1, 0.25, 5 and 10 microg). Conversely, intra-hippocampal infusions of R(+)-8-OH-DPAT (0.1 and 5 microg/side) produced substantial increases in open-arm activity in the plus-maze test, but did not alter rat burying behavior in the shock-probe test. These dissociations suggest that 5-HT1A receptors in the septum and hippocampus exert parallel but distinct control over different fear reactions.     

5-HT1A receptors activate a potassium conductance in rat ventromedial hypothalamic neurones.

The extracellularly recorded firing rate of rat ventromedial hypothalamic neurones in vitro was inhibited by 5-hydroxytryptamine (5-HT), 5-carboxamidotryptamine (5-CT), 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT) and buspirone, 2-methyl-5-HT was relatively ineffective. At 1 microM, spiperone and cyanopindolol antagonised the 5-CT induced inhibition, MDL 72222 (10 microM) and ketanserin (1 microM) did not. Intracellular recordings with voltage-clamp revealed that 5-HT and 5-CT evoked a tetrodotoxin-resistant outward current with a reversal potential of ca. -100 mV. 5-HT1A receptors likely activate a potassium conductance on these neurones.     

Evidence for excitatory 5-HT2-receptors on rat brainstem neurones.

1. The technique of microiontophoresis was used to investigate the identity of the receptor mediating the excitatory effects of 5-hydroxytryptamine (5-HT) upon neurones in the midline of the medullary brainstem of the rat in vivo. 2. The 5-HT1-like receptor agonists 5-carboxamidotryptamine (5-CT) and 8-hydroxy-2-(di-n-propylamino) tetralin (8-OH-DPAT) failed to excite the majority of neurones excited by 5-HT. The mobilities of 5-CT and 8-OH-DPAT when tested in vitro were found not to differ significantly from that of 5-HT, suggesting that the lack of effect of these agonists was not due to a lower rate of release from the microelectrodes. 3. The excitatory responses to 5-HT were attenuated by the 5-HT 2-receptor antagonists ketanserin and methysergide when applied microiontophoretically or administered intravenously (0.3 and 1 mg kg-1 respectively). Excitatory responses to glutamate and noradrenaline were not reduced. 4. The 5-HT3-receptor antagonist MDL 72222 failed to attenuate selectively the excitatory response to 5-HT when applied either by microiontophoresis or administered intravenously (1 mg kg-1). 5. Microiontophoretic application of the alpha 1-adrenoceptor antagonist prazosin did not attenuate excitatory responses to either 5-HT or noradrenaline. Intravenously administered prazosin (0.8 mg kg-1) also failed to attenuate excitatory responses to 5-HT, but did block excitatory responses to noradrenaline. 6. These results suggest that 5-HT2-receptors, but not 5-HT1-like receptors, 5-HT3-receptors or alpha 1-adrenoceptors, are involved in the excitatory response of midline medullary neurones to 5-HT.     

Possible in vivo 5-HT reuptake blocking properties of 8-OH-DPAT assessed by measuring hippocampal extracellular 5-HT using microdialysis in rats.

1. The 5-hydroxytryptamine (5-HT)1A receptor agonist, 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT), has been shown to label 5-HT reuptake sites. 2. To study the functional consequences of this property, the effects of 8-OH-DPAT were compared with those of the 5-HT reuptake inhibitors, paroxetine and clomipramine, and of the 5-HT1A receptor agonist flesinoxan, in vitro on 5-HT reuptake, and in vivo on the extracellular concentration of 5-HT by use of microdialysis, in rat hippocampus. Because 5-HT reuptake inhibitors reportedly attenuate the ability of (+)-fenfluramine to increase the extracellular concentration of 5-HT, the possible reversal of these effects of 8-OH-DPAT and by paroxetine were examined. 3. 8-OH-DPAT, paroxetine and clomipramine inhibited [3H]-5-HT reuptake in rat hippocampal synaptosomes (pIC50: 6.00, 8.41 and 7.00, respectively). In contrast, flesinoxan did not alter 5-HT reuptake (pIC50 < 5). 4. 8-OH-DPAT (10 and 100 microM), paroxetine (0.1 microM) and clomipramine (1 microM), administered through the dialysis probe, significantly increased the hippocampal extracellular concentration of 5-HT. In contrast, flesinoxan (100 microM) did not alter extracellular 5-HT. Moreover, the effects of 100 microM 8-OH-DPAT were not blocked by the 5-HT1A receptor antagonist, WAY-100635 (0.16 mg kg-1, s.c.). 5. The increase in extracellular 5-HT induced by 10 mg kg-1, i.p., (+)-fenfluramine was prevented not only by 0.1 microM paroxetine, but also by 100 microM 8-OH-DPAT. In addition, systemic administration of 10 mg kg-1, but not 2.5 mg kg-1, i.p. 8-OH-DPAT attenuated the increase in extracellular 5-HT induced by 2.5 mg kg-1, i.p., (+)-fenfluramine. 6. These findings suggest that the increase in extracellular 5-HT produced by local administration of 8-OH-DPAT does not involve its 5-HT1A receptor agonist properties, but may result, at least in part, from its 5-HT reuptake blocking properties.     

Serotonin-2 receptor-mediated regulation of release of acetylcholine by minaprine in cholinergic nerve terminal of hippocampus of rat

5-Hydroxytryptamine (5-HT) inhibited the K+-induced release of [3H]acetylcholine [( 3H]ACh) from slices of the hippocampus of the rat, dose-dependently. Minaprine (3-(2-morpholinoethylamino)-4-methyl-6-phenylpyridazine, Fig. 1) had no effect on the release of [3H]ACh. However, it inhibited the (formula; see text) Fig. 1. Chemical structure of minaprine dihydrochloride. attenuation of the release of [3H]ACh by 5-HT dose-dependently. The 5-HT2 receptor antagonists, mianserine, methysergide and spiperone, prevented the inhibitory effect of the 5-HT, as well as did minaprine. The attenuating effect of 5-HT was not mimicked by the 5-HT1A receptor agonist 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT) and was not prevented by a 5-HT1A and 5-HT1B mixed receptor antagonist, propranolol, or by the 5-HT3 receptor antagonists, cocaine and metoclopramide. Minaprine inhibited the bindings of [3H]5-HT, [3H]8-OH-DPAT and [3H]ketanserin in the hippocampus. The inhibitory effect of minaprine on the binding of [3H]ketanserin was more marked than on the binding of [3H]5-HT and [3H]8-OH-DPAT, and was non-competitive. The Ki value of minaprine for the binding of [3H]ketanserin was 2.9 microM. The inhibitory effect of 5-HT on the release of [3H]ACh was observed in the presence of tetrodotoxin. By electrolytic lesioning of the medial septum, the K+-induced release of [3H]ACh from the slices of hippocampus was significantly reduced and the release was no longer inhibited by 5-HT. The lesioning significantly decreased the binding of [3H]ketanserin in the hippocampus, with hardly any reduction in the binding of [3H]5-HT and [3H]8-OH-DPAT.(ABSTRACT TRUNCATED AT 250 WORDS)     

Block by capsaicin of voltage-gated K+ currents in melanotrophs of the rat pituitary.

1 The present study compares the effects on representative autonomic outflows of IVth ventricular application of tryptamine analogues which act at 5-HT1 receptors with 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT). 2 Cumulative doses of 8-OH-DPAT, N,N-di-n-propyl-5-carboxamidotryptamine (DP-5-CT) and 5-carboxamidotryptamine (5-CT, 2.5-40 nmol kg-1), sumatriptan (10-160 nmol kg-1), indorenate (100-800 nmol kg-1), 5-hydroxytryptamine (5-HT, 20-640 nmol kg-1) both alone and in the presence of cinanserin (0.1 mg kg-1) were given into the IVth ventricle of cats which were anaesthetized with a mixture of alpha-chloralose and pentobarbitone sodium, neuromuscularly blocked and artificially ventilated. Recordings were made of arterial blood pressure, heart rate, renal, cardiac, splanchnic and phrenic nerve activities, femoral arterial flow, tracheal and intragastric pressures. 3 Central application of each of the agonists evoked significant falls in arterial blood pressure. In addition 8-OH-DPAT, DP-5-CT, 5-CT and 5-HT all evoked a differential inhibition of sympathetic nerve activities, with renal nerve activity being the most sensitive and cardiac nerve activity the least sensitive. In the dose-ranges used, administration of sumatriptan evoked reductions only in renal and splanchnic nerve activities whilst indorenate reduced activity in all three sympathetic nerves to a similar extent. 4 The effect of the agonists on heart rate was more inconsistent than the effects on sympathetic outflow.(ABSTRACT TRUNCATED AT 250 WORDS)     

Evidence for depressant 5-HT1-like receptors on rat brainstem neurones.

1. The technique of microiontophoresis was used to evaluate the contribution of 5-HT1-like, 5-HT2- and 5-HT3-receptors to the depressant effects of 5-hydroxytryptamine (5-HT) on neurones in the midline of the medullary brainstem of the rat in vivo. 2. Depressant responses to 5-HT were resistant to antagonism by the 5-HT2-receptor antagonist ketanserin and the 5-HT3-receptor antagonist MDL 72222 applied either microiontophoretically or administered systemically. 3. Microiontophoretic or systemic administration of the 5-HT antagonist metergoline, which shows nanomolar affinity for the 5-HT1-binding site, also failed to attenuate the depressant responses to 5-HT. 4. Systemic administration of high doses of methysergide (30-40 mg kg-1) attenuated the depressant responses to 5-HT but did not block depressant responses to GABA or excitatory responses to glutamate. 5. The depressant effects of 5-HT were potently mimicked by the 5-HT1-like receptor agonists 5-carboxamidotryptamine and 8-OH-DPAT. 6. These results indicate that neither 5-HT2-receptors nor 5-HT3-receptors are involved in the depressant effects of 5-HT on midline brainstem neurones. The depressant effects of 5-carboxamidotryptamine (5-CT) and 8-hydroxy-2-(di-n-propylamino) tetralin (8-OH-DPAT) and blockade of the response to 5-HT by high doses of methysergide suggests the involvement of 5-HT1-like receptors. The lack of effect of metergoline, however, indicates that this receptor may be different from any of the 5-HT1 binding sites yet described.     

Inhibition of 5-carboxamidotryptamine-induced relaxation of guinea-pig ileum correlates with [125I]LSD binding

5-Hydroxytryptamine (5-HT), 5-carboxamidotryptamine (5-CT), 8-OH-DPAT and RU 24969 relax the guinea-pig ileum precontracted with histamine. This relaxation was characterized using 5-CT as agonist with a series of 8 competitive antagonists. [125I]LSD binding was measured in a membrane preparation of the longitudinal muscle of the guinea-pig ileum in the presence of 3 X 10(-7) M cinanserin in order to suppress binding to 5-HT2 receptors. There was a significant correlation (P less than 0.01) between the antagonism of 5-CT-induced relaxation (pA2 values) and the affinity values of the antagonists for [125I]LSD binding (pKD values). It was also shown that 5-CT relaxed histamine-precontracted longitudinal muscle strips of the guinea-pig ileum. The results suggest that the 5-HT receptor-mediating relaxation in the guinea-pig ileum can be labelled with [125I]LSD and that this receptor does not belong to the 5-HT2, 5-HT1A, 5-HT1B or 5-HT1C receptor subtypes.     

Role of the medial prefrontal cortex in 5-HT1A receptor-induced inhibition of 5-HT neuronal activity in the rat

1. We examined the involvement of the frontal cortex in the 5-HT2A receptor-induced inhibition of 5-HT neurones in the dorsal raphe nucleus (DRN) of the anaesthetized rat using single-unit recordings complemented by Fos-immunocytochemistry. 2. Both transection of the frontal cortex as well as ablation of the medial region of the prefrontal cortex (mPFC) significantly attenuated the inhibition of 5-HT neurones induced by systemic administration of the 5-HT1A receptor agonist, 8-OH-DPAT (0.5-16 microg kg(-1), i.v.). In comparison, the response to 8-OH-DPAT was not altered by ablation of the parietal cortex. The inhibitory effect of 8-OH-DPAT was reversed by the 5-HT1A receptor antagonist, WAY 100635 (0.1 mg kg(-1), i.v.) in all neurones tested. 3. In contrast, cortical transection did not alter the sensitivity of 5-HT neurones to iontophoretic application of 8-OH-DPAT into the DRN. Similarly, cortical transection did not alter the sensitivity of 5-HT neurones to systemic administration of the selective 5-HT reuptake inhibitor, paroxetine (0.1-0.8 mg kg(-1) , i.v.). 4. 8-OH-DPAT evoked excitation of mPFC neurones at doses (0.5-32 microg kg(-1), i.v.) in the range of those which inhibited 5-HT cell firing. At higher doses (32-512 microg kg(-1), i.v.) 8-OH-DPAT inhibited mPFC neurones. 8-OH-DPAT (0.1 mg kg(-1), s.c.) also induced Fos expression in the mPFC. The neuronal excitation and inhibition, as well as the Fos expression, were antagonized by WAY 100635. 5. These data add further support to the view that the inhibitory effect of 5-HT1A receptor agonists on the firing activity of DRN 5-HT neurones involves, in part, activation of a 5-HT1A receptor-mediated postsynaptic feedback loop centred on the mPFC.     

Increased dopamine turnover in the ventral striatum by 8-OH-DPAT administration in the rat

The administration of the 5-HT1A agonist 8-OH-DPAT (0.8 mumols kg-1 s.c.-40 min) produced an increase in dopamine (DA) turnover, estimated by the quotient (DOPAC + HVA) DA-1, in the ventral striatum of the rat. No statistically significant effects were obtained in the dorsal striatum. The accumulation of 3-MT in pargyline-treated animals (375 mumols kg-1 s.c.-60 min) was not affected by 8-OH-DPAT treatment (0.15-2.4 mumols kg-1 s.c.-30 min). These findings indicate that 8-OH-DPAT has weak antagonist properties at striatal DA receptors in normal rats. Both the 5-HT1A agonist flesinoxan (0.06-17.8 mumos kg-1 s.c. -50 min) and the mixed 5-HT1 and 5-HT2 agonist 5-MeODMT (1.6-26.0 mumols kg-1 s.c.-50 min) produced a decrease in forebrain 5-HTP accumulation (striatum and neocortex), following decarboxylase inhibition by means of NSD-1015 in reserpine treated rats, indicating stimulation of central 5-HT receptors by these two compounds. At the same time, the DOPA accumulation by the ventral striatum was decreased by flesinoxan and increased by 5-MeODMT treatment. These observations show that, under these conditions, the decrease in DA synthesis is not directly coupled to the decreased 5-HT synthesis produced by flesinoxan, as previously demonstrated for 8-OH-DPAT. Taken together with previous observations, the present results suggest that 8-OH-DPAT, depending on the experimental conditions, is an agonist or antagonist at striatal DA receptors, in all probability due to partial DA receptor agonist properties of the compound.