Identification of biomarkers for glycaemic deterioration in type 2 diabetes.

We identify biomarkers for disease progression in three type 2 diabetes cohorts encompassing 2,973 individuals across three molecular classes, metabolites, lipids and proteins. Homocitrulline, isoleucine and 2-aminoadipic acid, eight triacylglycerol species, and lowered sphingomyelin 42:2;2 levels are predictive of faster progression towards insulin requirement. Of ~1,300 proteins examined in two cohorts, levels of GDF15/MIC-1, IL-18Ra, CRELD1, NogoR, FAS, and ENPP7 are associated with faster progression, whilst SMAC/DIABLO, SPOCK1 and HEMK2 predict lower progression rates. In an external replication, proteins and lipids are associated with diabetes incidence and prevalence. NogoR/RTN4R injection improved glucose tolerance in high fat-fed male mice but impaired it in male db/db mice. High NogoR levels led to islet cell apoptosis, and IL-18R antagonised inflammatory IL-18 signalling towards nuclear factor kappa-B in vitro. This comprehensive, multi-disciplinary approach thus identifies biomarkers with potential prognostic utility, provides evidence for possible disease mechanisms, and identifies potential therapeutic avenues to slow diabetes progression.

Intra-islet glucagon confers ß-cell glucose competence for first-phase insulin secretion and favors GLP-1R stimulation by exogenous glucagon.

We report that intra-islet glucagon secreted from α-cells signals through ß-cell glucagon and GLP-1 receptors (GcgR and GLP-1R), thereby conferring to rat islets their competence to exhibit first-phase glucose-stimulated insulin secretion (GSIS). Thus, in islets not treated with exogenous glucagon or GLP-1, first-phase GSIS is abolished by a GcgR antagonist (LY2786890) or a GLP-1R antagonist (Ex[9-39]). Mechanistically, glucose competence in response to intra-islet glucagon is conditional on ß-cell cAMP signaling because it is blocked by the cAMP antagonist prodrug Rp-8-Br-cAMPS-pAB. In its role as a paracrine hormone, intra-islet glucagon binds with high affinity to the GcgR, while also exerting a "spillover" effect to bind with low affinity to the GLP-1R. This produces a right shift of the concentration-response relationship for the potentiation of GSIS by exogenous glucagon. Thus, 0.3 nM glucagon fails to potentiate GSIS, as expected if similar concentrations of intra-islet glucagon already occupy the GcgR. However, 10 to 30 nM glucagon effectively engages the ß-cell GLP-1R to potentiate GSIS, an action blocked by Ex[9-39] but not LY2786890. Finally, we report that the action of intra-islet glucagon to support insulin secretion requires a step-wise increase of glucose concentration to trigger first-phase GSIS. It is not measurable when GSIS is stimulated by a gradient of increasing glucose concentrations, as occurs during an oral glucose tolerance test in vivo. Collectively, such findings are understandable if defective intra-islet glucagon action contributes to the characteristic loss of first-phase GSIS in an intravenous glucose tolerance test that is diagnostic of type 2 diabetes in the clinical setting.

Episodic ß-cell death and dedifferentiation during diet-induced obesity and dysglycemia in male mice.

Loss of functional islet ß-cell mass through cellular death or dedifferentiation is thought to lead to dysglycemia during the progression from obesity to type 2 diabetes. To assess these processes in a mouse model of obesity, we performed measures of circulating cell-free differentially methylated insulin II ( Ins2) DNA as a biomarker of ß-cell death and aldehyde dehydrogenase 1 family member A3 (ALDH1A3) and forkhead box 01 (Foxo1) immunostaining as markers of ß-cell dedifferentiation. Eight-week-old, C57BL/6J mice were fed a low-fat diet (LFD; 10% kcal from fat) or a high-fat diet (HFD; 60% kcal from fat) and were followed longitudinally for up to 13 wk to measure glycemic control and ß-cell mass, death, and dedifferentiation. Compared with LFD controls, ß-cell mass increased during the feeding period in HFD animals, and statistically greater ß-cell death (unmethylated Ins2) was detectable at 2 and 6 wk after diet initiation. Those times correspond to periods when significant step increases in fasting glucose and glucose intolerance, respectively, were detected. ALDH1A3 and Foxo1 immunostaining of the pancreas revealed evidence of ß-cell dedifferentiation by 13 wk when fed an HFD, but not in LFD controls. In conclusion, early episodic ß-cell death may be a feature of cellular turnover correlated with changes in glycemia during ß-cell mass accrual in obesity, whereas ß-cell dedifferentiation may be a feature seen later in established disease.-Tersey, S. A., Levasseur, E. M., Syed, F., Farb, T. B., Orr, K. S., Nelson, J. B., Shaw, J. L., Bokvist, K., Mather, K. J., Mirmira, R. G. Episodic ß-cell death and dedifferentiation during diet-induced obesity and dysglycemia in male mice.

5-Hydroxytryptamine 1A (5HT1A) receptors mediate increases in plasma glucose independent of corticosterone.

Hypothalamic 5HT1A receptors play an important role in the regulation of satiety, glycemia and endocrine status. In the present study, 8-OH-DP administered centrally and peripherally to C57/Bl6 mice and plasma glucose insulin and corticosterone were evaluated. In these studies, dose and time dependent increases in glucose and corticosterone were observed while no alterations in insulin were seen. The increases in plasma corticosterone were prevented by prior central or peripheral administration of LY426965, a specific 5HT1A antagonist. Intracerebroventricular coadministration of a 5HT1A antagonist with 8-OH-DPAT prevented the increase in plasma glucose establishing this response as a centrally mediated response in mice. Given that increases in plasma corticosterone are associated with increases in plasma glucose, we conducted experiments to determine if increased plasma corticosterone was the mechanism by which 8-OH-DPAT increased plasma glucose. Prior administration of the glucocorticoid antagonist mifepristone did not affect the increase in plasma glucose produced by 8-OH-DPAT. Prior administration of the glucocorticoid synthesis inhibitor, metyrapone, reduced basal corticosterone and the concentrations of corticosterone associated with 8-OH-DPAT administration. However, metyrapone administration did not affect the increases in plasma glucose. Therefore, 5HT1A receptors regulate glucose through brain mechanisms, but not through regulation of the hypophyseal-pituitary axis. Antagonism of brain 5HT1A receptors may enable discovery of novel antidiabetic agents.

LLY-2707, a novel nonsteroidal glucocorticoid antagonist that reduces atypical antipsychotic-associated weight gain in rats.

Weight gain and diabetes have been reported during treatment with atypical antipsychotic drugs (AAPDs). Patients treated with the glucocorticoid receptor antagonist (GRA) and the progesterone receptor antagonist (PRA) mifepristone [estra-4,9-dien-3-one, 11-[4-(dimethylamino)phenyl]-17-hydroxy-17-(1-propynyl)-(11ß,17ß)-(9CI)] experienced significant reduction in the weight gain observed when patients were treated with olanzapine or risperidone. To understand the pharmacology responsible for this finding, we discovered LLY-2707 [N-(5-(tert-butyl)-3-(2-fluoro-5-methylpyridin-4-yl)-2-methyl-1H-indol-7-yl)methanesulfonamide], a novel and selective GRA, and evaluated its utility in preclinical models of AAPD-associated weight gain and diabetes. In vitro, LLY-2707 was a highly selective and potent GRA. GR occupancy in vivo was assessed using ex vivo binding where LLY-2707 inhibited [(3)H]dexamethasone binding to the liver. Modest but statistically significant decreases in brain ex vivo binding were observed with high doses of CORT-108297 [(R)-4α-(ethoxymethyl)-1-(4-fluorophenyl)-6-((4-(trifluoromethyl)phenyl)sulfonyl)-4,4a,5,6,7,8-hexahydro-1H-pyrazolo[3,4-g]isoquinoline] and LLY-2707, but mifepristone inhibited at all doses. Central activity of the GRAs was confirmed by their ability to suppress amphetamine-induced increases in locomotor activity. The increases in the body weight of female rats treated with olanzapine (2 mg/kg PO) over 14 days were reduced in a dose-dependent manner by coadministration of LLY-2707. Similar decreases, although less robust, in body weight were seen with mifepristone and CORT-108297. In addition, sGRAs prevented the glucose excursion after intragastric olanzapine infusions consistent with a direct effect on the hyperglycemia observed during treatment with AAPDs. At doses effectively preventing weight gain, LLY-2707 did not substantially interfere with the dopamine D2 receptor occupancy by olanzapine. Therefore, GRA coadministration may provide a novel treatment modality to prevent the weight gain and diabetes observed during treatment with AAPDs.

5-HT1A receptor antagonists reduce food intake and body weight by reducing total meals with no conditioned taste aversion.

Serotonin acts through receptors controlling several physiological functions, including energy homeostasis regulation and food intake. Recent experiments demonstrated that 5-HT1A receptor antagonists reduce food intake. We sought to examine the microstructure of feeding with 5-HT1A receptor antagonists using a food intake monitoring system. We also examined the relationship between food intake, inhibition of binding and pharmacokinetic (PK) profiles of the antagonists. Ex vivo binding revealed that, at doses used in this study to reduce food intake, inhibition of binding of a 5-HT1A agonist by ~40% was reached in diet-induced obese (DIO) mice with a trend for higher binding in DIO vs. lean animals. Additionally, PK analysis detected levels from 2 to 24h post-compound administration. Male DIO mice were administered 5-HT1A receptor antagonists LY439934 (10 or 30 mg/kg, p.o.), WAY100635 (3 or 10mg/kg, s.c.), SRA-333 (10 or 30 mg/kg, p.o.), or NAD-299 (3 or 10mg/kg, s.c.) for 3 days and meal patterns were measured. Analyses revealed that for each antagonist, 24-h food intake was reduced through a specific decrease in the total number of meals. Compared to controls, meal number was decreased 14-35% in the high dose. Average meal size was not changed by any of the compounds. The reduction in food intake reduced body weight 1-4% compared to Vehicle controls. Subsequently, a conditioned taste aversion (CTA) assay was used to determine whether the feeding decrease might be an indicator of aversion, nausea, or visceral illness caused by the antagonists. Using a two bottle preference test, it was found that none of the compounds produced a CTA. The decrease in food intake does not appear to be a response to nausea or malaise. These results indicate that 5-HT1A receptor antagonist suppresses feeding, specifically by decreasing the number of meals, and induce weight loss without an aversive side effect.

Suppression of alcohol self-administration and reinstatement of alcohol seeking by melanin-concentrating hormone receptor 1 (MCH1-R) antagonism in Wistar rats.

RATIONALE: Melanin-concentrating hormone (MCH) is involved in regulation of appetitive behaviors as well as emotional reactivity and reward, behavioral domains relevant to alcohol addiction. MATERIALS AND METHODS: We evaluated the effects of the non-peptide MCH1 receptor antagonist, GW803430 [6-(4-chloro-phenyl)-3-[3-methoxy-4-(2-pyrrolidin-1-yl-ethoxy)-phenyl]-3H-thieno[3,2-d]pyrimidin-4-one; 3-30 mg/kg, i.p.] on alcohol-related behaviors in Wistar rats. RESULTS: Ex vivo binding experiments demonstrated that the GW803430 dose range used resulted in high central MCH1 receptor occupancy. Alcohol self-administration was dose-dependently and potently suppressed, by approximately 80% at the highest dose. Reinstatement of alcohol-seeking induced by alcohol-associated cues was essentially eliminated. In contrast, reinstatement induced by footshock stress was not significantly altered. Taste preference for a quinine/saccharin solution, locomotor activity, and alcohol elimination were unaffected. CONCLUSION: Together, these observations support a specific involvement of the MCH system in mediating alcohol reward and cue-induced relapse to alcohol seeking. MCH1-R antagonism may constitute an attractive treatment target for alcohol use disorders.

Preclinical evaluation of melanin-concentrating hormone receptor 1 antagonism for the treatment of obesity and depression.

The mammalian neuropeptide, melanin-concentrating hormone, interacts with two G protein-coupled receptors, melanin-concentrating hormone receptor (MCHR) 1 and MCHR2; however, only MCHR1 is expressed in rats and mice. In the present study, we evaluated MCHR1 antagonism in preclinical models believed to be predictive of antiobesity and antidepressant activity. Central activity of the selective MCHR1 antagonist, GW803430 [6-(4-chloro-phenyl)-3-[3-methoxy-4-(2-pyrrolidin-1-yl-ethoxy)-phenyl]-3H-thieno[3,2-d]pyrimidin-4-one], was evaluated using ex vivo binding with autoradiography. Effective doses of GW803430 (1 and 3 mg/kg p.o.) were correlated with antiobesity activity in a 14-day study of diet-induced obese rats. GW803430 was evaluated subsequently for antidepressant-like effects in mice and rats. Acute and subchronic administration reduced immobility time in the mouse forced-swim test at doses of 3 (acute) and 3 and 10 (chronic) mg/kg p.o., an effect that was absent in MCHR1(-/-) mice. Combined subeffective doses of GW803430 (0.3 and 1 mg/kg p.o.) and imipramine (5 mg/kg) produced a robust antidepressant-like response. The compound was also active in the tail suspension test at a dose of 10 mg/kg p.o. GW803430 (30 mg/kg p.o.) significantly reduced submissive behaviors at weeks 2 and 3, a model of submissive behavior that may predict antidepressant onset. GW803430 decreased marble burying in mice at doses of 3, 10, and 30 mg/kg p.o., an assay that detects anxiolytic-like effects. Thus, GW803430 produces robust antiobesity and antidepressant-like effects in rats and mice at doses that compete for central MCHR1 in vivo. As such, MCHR1 should be considered as a promising target for future drug discovery efforts.

Monoaminergic compensation in the neuropeptide Y deficient mouse brain.

Neuropeptide Y (NPY) is an important central regulator of food consumption and energy expenditure via the hypothalamus. NPY containing neurons have a broad central distribution and are often colocalized with norepinephrine (NE). However, NPY deficient mice do not exhibit any substantial changes in food consumption, body weight or body composition when compared to wild type mice. Since NE and serotonin (5HT) are also important regulators of appetite and metabolism, we evaluated these systems in NPY deficient mice. Brain sections from NPY deficient and wild type mice were labeled with either (3)H-nisoxetine for the NE transporter (NET) or (3)H-citalopram for the 5HT transporter (SERT). Tyrosine hydroxylase expression was evaluated by radioimmunohistochemistry. Brain monoamines and metabolites were evaluated using HPLC. NPY deficient mice exhibited a substantial decrease in NET binding in most brain regions examined. NET binding was less than 50% of control binding in the cerebral cortex and subregions of the thalamus with the greatest decrease seen in the hypothalamus. In contrast, more modest and regionally variable changes were observed in the SERT binding with decreases in regions such as the accessory olfactory nucleus, glomerular layer of the olfactory bulb and the CA1 region of the hippocampus. Measurement of NE and 5HT content as well as the primary metabolites revealed increased NE turnover and decreased 5HT content in the hypothalamus. Therefore, developmental compensation by the NE and 5HT systems may contribute to the absence of a body weight phenotype in NPY deficient mice.

Increased brain neuropeptide Y1 and Y2 receptor binding in NPY knock out mice does not result in increased receptor function.

The brain neuropeptide Neuropeptide Y (NPY) is an important modulator of a number of centrally mediated processes including feeding, anxiety-like behaviors, blood pressure and others. NPY produces its effects through at least four functional G-protein coupled receptors termed Y1, Y2, Y4 and Y5. In the brain, the Y1 and Y2 receptor subtypes are the predominant receptor population. To better understand the roles of NPY, genetically modified mice lacking NPY were produced but lacked the expected phenotypes. These mice have previously been reported to have a marked increase in Y2 receptor binding. In the present study, we found an upregulation of both Y1 and Y2 receptor binding and extended these findings to the female. These increases were as large as 10-fold or greater in many brain regions. To assess functional coupling of the receptors, we performed agonist-induced [(35)S]GTPgammaS autoradiography. In the mouse brain, the Y1/Y4/Y5 agonist Leu(31),Pro(34)-NPY increased [(35)S]GTPgammaS binding with a regional distribution consistent with that produced when labeling adjacent sections with [(125)I]-Leu(31),Pro(34)-PYY. In a few brain regions, minor increases were noted in the agonist-induced binding when comparing knock out mice to wild type. The Y2 agonist C2-NPY stimulated [(35)S]GTPgammaS binding in numerous brain areas with a regional distribution similar to the binding observed with [(125)I]-PYY3-36. Again, no major increases were noted in the functional activation of Y2 receptors between knock out and wild type mice. Therefore, the increased Y1 and Y2 binding observed in the NPY knock out mice does not represent an increase in NPY receptor mediated signaling and is likely due to an increase in spare (uncoupled) receptors.

Distribution of nociceptin and Ro64-6198 activated [35S]-GTPgammaS binding in the rat brain.

The peptide, nociceptin, was discovered as the endogenous ligand for the opioid-like receptor, ORL1. Since its discovery, this peptide has been shown to modulate the perception of pain, modulate feeding and produce behavioral effects in rodent models of mood disorders. Recently, the non-peptide agonist {(1S,3aS)-8-(2,3,3a,4,5,6-hexahydro-1H-phenalen-1-yl)-1-phenyl-1,3,8-triaza-spiro[4,5]decan-4-one} (Ro64-6198) of the ORL1 receptor has been reported in the literature. In the present study, we compared the distribution and potency of Ro64-6198 with nociceptin for their ability to stimulate [(35)S]-GTPgammaS binding to sections of rat brain. In initial studies, Ro64-6198 inhibited (125)I-nociceptin binding to the hORL1 receptors with a K(i) of 1.75 nM compared with 0.20 nM for nociceptin. To assess agonist potency in a whole cell assay, a cell line expressing the hORL1 receptor and G(alpha15) was created and used for calcium mobilization studies. In this assay system, Ro64-6198 increased intracellular calcium with an EC(50) of 52nM compared with 24 nM for nociceptin. Having verified the agonist properties of Ro64-6198, we then assessed the potency and distribution of ORL1 receptor activation in rat brain sections. In dose-response studies, Ro64-6198 increased [(35)S]-GTPgammaS binding to a variety of brain regions with EC(50) values ranging from 84.9 to 2,143 nM depending on the brain regions evaluated. These potencies were similar to that seen for nociceptin, but substantially lower than values established using [(125)I] nociceptin binding to the cloned human ORL1 receptor. In general, the brain distribution of agonist stimulated [(35)S]-GTPgammaS binding was similar when either Ro64-6198 or nociceptin were used. Using these techniques, we have demonstrated, for the first time that Ro64-6198 activates [(35)S]-GTPgammaS binding to rat brain sections and this compound stimulates a similar population of receptors as nociceptin.

Advances toward new antidepressants beyond SSRIs: 1-aryloxy-3-piperidinylpropan-2-ols with dual 5-HT1A receptor antagonism/SSRI activities. Part 5.

A series of 1-aryloxy-3-piperidinylpropan-2-ols possessing potent dual 5-HT1A receptor antagonism and serotonin reuptake inhibition was discovered. 1-(1H-Indol-4-yloxy)-3-(4-benzo[b]thiophen-2-ylpiperidinyl)propan-2-ols exhibited selective and high affinities at the 5-HT1A receptor and serotonin reuptake site in vitro. In vivo evaluation of this series of compounds demonstrated elevated extracellular serotonin levels from the basal and quick recovery of neuron firing that was presumably suppressed by the initial acute activation of 5-HT1A somatodendritic autoreceptors.

Stress and central Urocortin increase anxiety-like behavior in the social interaction test via the CRF1 receptor.

Corticotropin releasing factor (CRF) and Urocortin are important neurotransmitters in the regulation of physiological and behavioral responses to stress. Centrally administered CRF or Urocortin produces anxiety-like responses in numerous animal models of anxiety disorders. Previous studies in our lab have shown that Urocortin infused into the basolateral nucleus of the amygdala produces anxiety-like responses in the social interaction test. Subsequently, in the current study we prepared a specific CRF1 receptor antagonist (N-Cyclopropylmethyl-2,5-dimethyl-N-propyl-N'-(2,4,6-trichloro-phenyl)-pyrimidine-4,6-diamine, NBI3b1996) to examine in this paradigm. This CRF1 receptor antagonist inhibited the ex vivo binding of 125I-sauvagine to rat cerebellum with an ED50 of 6 mg/kg, i.p. NBI3b1996 produced a dose-dependent antagonism of Urocortin-induced anxiety-like behavior in Social Interaction test with an ED50 of 6 mg/kg, i.p. The compound had no effect on baseline social interaction. In addition, the CRF1 receptor antagonist prevented the stress-induced decrease in social interaction. These results provide further support for the CRF1 receptor in anxiety-like behavior and suggest this pathway is quiescent in unstressed animals.

Mesolimbic dopamine super-sensitivity in melanin-concentrating hormone-1 receptor-deficient mice.

Melanin-concentrating hormone (MCH) neurons and MCH-1 receptors (MCH1r) densely populate mesolimbic dopaminergic brain regions such as the nucleus accumbens (NAc). The regulation of dopamine by MCH1r was suggested to be an important mechanism underlying the hyperactive phenotype of MCH1r knock-out (ko) mice. However, MCH1r modulation of monoamine neurotransmission has yet to be examined. We tested whether dopamine, norepinephrine, and serotonin function is dysregulated in MCH1r ko and wild-type (wt) mice. MCH1r ko mice exhibited robust hyperactivity in a novel or familiar environment and were super-sensitive to the locomotor activating effects of d-amphetamine and the D1 agonist 2,3,4,5-tetrahydro-7,8-dihydroxy-1-phenyl-1H-3-benazepine HCl. The D2 agonist, quinpirole, decreased locomotion similarly in both ko and wt mice. Tissue contents of dopamine within the NAc and caudate-putamen were not significantly different in ko compared with wt mice. Basal and amphetamine-evoked NAc dopamine, norepinephrine, and serotonin efflux, as measured using in vivo microdialysis, were not significantly different between genotypes. In contrast, D1-like and D2-like receptor binding were significantly higher within the olfactory tubercle, ventral tegmental area, and NAc core and shell of ko mice. Norepinephrine transporter (NET) binding was significantly elevated within the NAc shell and globus pallidus of ko mice, whereas serotonin transporter binding was decreased in the NAc shell. Thus, deletion of MCH1r results in an upregulation of mesolimbic dopamine receptors and NET, indicating that MCH1r may negatively modulate mesolimbic monoamine function. MCH1r may be an important therapeutic target for neuropsychiatric disorders involving dysregulation of limbic monoamine systems.

Advances toward new antidepressants beyond SSRIs: 1-aryloxy-3-piperidinylpropan-2-ols with dual 5-HT1A receptor antagonism/SSRI activities. Part 4.

A series of 1-(1H-indol-4-yloxy)-3-(4-arylpiperidinyl)propan-2-ols possessing potent dual 5-HT(1A) receptor antagonism and serotonin reuptake inhibition was discovered. The fused aryl ring moiety contributed to the robust dual activities irrespective of the regiochemistry associated with its connectivity to the piperidine central ring.

Functional autoradiography of neuropeptide Y Y1 and Y2 receptor subtypes in rat brain using agonist stimulated [35S]GTPgammaS binding.

Neuropeptide Y, one of the most abundant brain peptides, has been found to modulate several important biological functions via a family of G-protein coupled receptors. To investigate the localization of functional NPY receptor subtypes in the rat brain, we performed agonist-induced [35S]GTPgammaS autoradiography. The Y1/Y4/Y5 agonist Leu(31), Pro(34)-NPY increased [35S]GTPgammaS binding in several brain areas with a regional distribution consistent with that produced when labeling adjacent sections with [125I]-Leu(31), Pro(34)-PYY. The Y1 selective antagonist BIBP3226 antagonized the Leu(31), Pro(34)-NPY stimulated increase in [35S]GTPgammaS binding in all areas examined. The Y2 agonist C2-NPY stimulated [35S]GTPgamma binding in numerous brain areas with a regional distribution similar to the binding observed with [125I]-PYY 3-36. No increase in [35S]GTPgammaS binding above basal was observed in any brain area evaluated using Y4 and Y5 selective agonists. This study demonstrates abundant Y1 and Y2 receptor activation in the rat brain, while evidence for functional Y4 and Y5 receptors was not observed.

Advances toward new antidepressants beyond SSRIs: 1-aryloxy-3-piperidinylpropan-2-ols with dual 5-HT1A receptor antagonism/SSRI activities. Part 3.

A series of 1-aryloxy-3-piperidinylpropan-2-ols possessing potent dual 5-HT(1A) receptor antagonism and serotonin reuptake inhibition was discovered. Modification of potential metabolic sites of 1-(1H-indol-4-yloxy)-3-(4-benzo[b]thiophen-2-ylpiperidinyl)propan-2-ols further improved the in vitro binding affinities and functional antagonism.

Advances Toward new antidepressants beyond SSRIs: 1-aryloxy-3-piperidinylpropan-2-ols with dual 5-HT(1A) receptor antagonism/SSRI activities. Part 2.

Potent 5-HT1A/SSRIs at low nanomolar and subnanomolar concentrations were identified in a series of 1-(1H-indol-4-yloxy)-3-(4-benzo[b]thiophen-2-ylpiperidinyl)propan-2-ols. Incorporation of an alpha-Me group in the piperidine ring with its specific stereochemistry enhanced binding affinity at the 5-HT reuptake site and in vitro 5-HT(1A) antagonist functional activity.

Advances toward new antidepressants beyond SSRIs: 1-aryloxy-3-piperidinylpropan-2-ols with dual 5-HT1A receptor antagonism/SSRI activities. Part 1.

A series of 1-aryloxy-3-piperidinylpropan-2-ols possessing potent dual 5-HT(1A) receptor antagonism and serotonin reuptake inhibition was discovered. 1-(1H-Indol-4-yloxy)-3-(4-benzo[b]thiophen-2-ylpiperidinyl)propan-2-ols exhibited selective and high affinity at the 5-HT(1A) receptor and serotonin reuptake inhibition at nanomolar concentrations for dual activities.

Fluoxetine, but not other selective serotonin uptake inhibitors, increases norepinephrine and dopamine extracellular levels in prefrontal cortex.

RATIONALE: The selective serotonin uptake inhibitor (SSRI) fluoxetine has been shown to not only increase the extracellular concentrations of serotonin, but also dopamine and norepinephrine extracellular concentrations in rat prefrontal cortex. The effect of other SSRIs on monoamine concentrations in prefrontal cortex has not been thoroughly studied. OBJECTIVE: The aim of this study was to compare the ability of five systemically administered selective serotonin uptake inhibitors to increase acutely the extracellular concentrations of serotonin, norepinephrine and dopamine in rat prefrontal cortex. METHODS: The extracellular concentrations of monoamines were determined in the prefrontal cortex of conscious rats using the microdialysis technique. RESULTS: Fluoxetine, citalopram, fluvoxamine, paroxetine and sertraline similarly increased the extracellular concentrations of serotonin from 2- to 4-fold above baseline. However, only fluoxetine produced robust and sustained increases in extracellular concentrations of norepinephrine and dopamine after acute systemic administration. Fluoxetine at the same dose blocked ex vivo binding to the serotonin transporter, but not the norepinephrine transporter, suggesting that the increase of catecholamines was not due to non-selective blockade of norepinephrine uptake. Prefrontal cortex extracellular concentrations of fluoxetine at the dose that increased extracellular monoamines were 242 nM, a concentration sufficient to block 5-HT(2C) receptors which is a potential mechanism for the fluoxetine-induced increase in catecholamines. CONCLUSION: Amongst the SSRIs examined, only fluoxetine acutely increases extracellular concentrations of norepinephrine and dopamine as well as serotonin in prefrontal cortex, suggesting that fluoxetine is an atypical SSRI.