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Chronic fluoxetine treatment selectively uncouples raphe 5-HT(1A) receptors as measured by [(35)S]-GTP gamma S autoradiography.

ABSTRACT: 1. Selective Serotonin Reuptake Inhibitors (SSRIs) are thought to have a delay in therapeutic efficacy because of the need to overcome the inhibitory influence of raphe 5-HT(1A) autoreceptors. Prolonged SSRI administration has been reported to desensitize these autoreceptors. We have used [(35)S]-GTP gamma S autoradiography to determine whether this desensitization occurs at the level of receptor/G protein coupling. 2. Male mice were injected intraperitoneally once a day with saline or 20 mg kg(-1) fluoxetine for either 2 days or 14 days. 5-HT(1A) receptor binding and coupling to G proteins were assessed using [(3)H]-8-OH-DPAT and [(35)S]-GTP gamma S autoradiography, respectively. 3. The 5-HT receptor agonist 5-carboxamidotryptamine (5-CT) stimulated [(35)S]-GTP gamma S binding in the substantia nigra, as well as in hippocampus and dorsal raphe nucleus. The 5-HT(1A) receptor antagonist p-MPPF (4-fluoro-N-(2-[4-(2-methoxyphenyl)1-piperazinyl]ethyl)-N-(2-pyridinyl)benzamide) blocked this effect in the latter regions, whereas the 5-HT(1B/D) antagonist GR-127,935 (2'-methyl-4'-(5-methyl-[1,2,4]oxadiazol-3-yl)-biphenyl-4-carboxylic acid [4-methoxy-3-(4-methyl-piperazin-l-yl)-phenyl]-amide) only decreased labelling in substantia nigra. 4. Fourteen-day fluoxetine treatment decreased 5-CT-stimulated [(35)S]-GTP gamma S binding in dorsal raphe (saline: 112 +/- 12% stimulation; fluoxetine: 66 +/- 13%), but not in substantia nigra (99 +/- 14% vs 103 +/- 7%) or hippocampus (157 +/- 3% vs 148 +/- 18%). Two-day fluoxetine treatment did not alter 5-CT-stimulated [(35)S]-GTP gamma S binding in any of the brain areas investigated. 5. Decreased [(35)S]-GTP gamma S binding was not due to receptor down-regulation, since the density of raphe [(3)H]-8-OH-DPAT binding sites was unaffected by fluoxetine treatment. 6. These results suggest that the desensitization of presynaptic 5-HT(1A) receptor function occurs at the level of receptor-G protein interaction on dorsal raphe neurons, and may underlie the therapeutic efficacy of long-term SSRI treatment.

SUBMITTER: Pejchal T

PROVIDER: S-EPMC1573219 | biostudies-other | 2002 Mar

REPOSITORIES: biostudies-other

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Project description:A major problem with current anti-depressant therapy is that it takes on average 6-7 weeks for remission. Since desensitization of serotonin (5-HT)1A receptor signaling contributes to the anti-depressive response, acceleration of the desensitization may reduce this delay in response to antidepressants. The purpose of the present study was to test the hypothesis that estradiol accelerates fluoxetine-induced desensitization of 5-HT1A receptor signaling in the paraventricular nucleus of the hypothalamus (PVN) of rats, via alterations in components of the 5-HT1A receptor signaling pathway. Ovariectomized rats were injected with estradiol and/or fluoxetine, then adrenocorticotropic hormone (ACTH) and oxytocin responses to a 5-HT1A receptor agonist (+)-8-hydroxy-2-dipropylaminotetralin (8-OH-DPAT) were examined to assess the function of 5-HT1A receptors in the PVN. Treatment with estradiol for either 2 or 7 days or fluoxetine for 2 days produced at most a partial desensitization of 5-HT1A receptor signaling, whereas 7 days of fluoxetine produced full desensitization. Combined treatment with estradiol and fluoxetine for 2 days produced nearly a full desensitization, demonstrating an accelerated response compared to either treatment alone. With two days of combined treatments, estradiol prevented the fluoxetine-induced increase in 5-HT1A receptor protein, which could contribute to the more rapid desensitization. Furthermore, EB treatment for 2 days decreased the abundance of the 35 kD G?z protein which could contribute to the desensitization response. We found two isoforms of G?z proteins with molecular mass of 35 and 33 kD, which differentially distributed in the detergent resistant microdomain (DRM) and in Triton X-100 soluble membrane region, respectively. The 35 kD G?z proteins in the DRM can be sumoylated by SUMO1. Stimulation of 5-HT1A receptors with 8-OH-DPAT increases the sumoylation of G?z proteins and reduces the 33 kD G?z proteins, suggesting that these responses may be related to the desensitization of 5-HT1A receptors. Treatment with estradiol for 2 days also reduced the levels of the G-protein coupled estrogen receptor GPR30, possibly limiting to the ability of estradiol to produce only a partial desensitization response. These data provide evidence that estradiol may be effective as a short-term adjuvant to SSRIs to accelerate the onset of therapeutic effects.

Project description:The ciliated protozoan Tetrahymena thermophila contains two distinct nuclei within a single cell-the mitotic micronucleus and the amitotic macronucleus. Although microtubules are required for proper division of both nuclei, macronuclear chromosomes lack centromeres and the role of microtubules in macronuclear division has not been established. Here we describe nuclear division defects in cells expressing a mutant beta-tubulin allele that confers hypersensitivity to the microtubule-stabilizing drug paclitaxel. Macronuclear division is profoundly affected by the btu1-1 (K350M) mutation, producing cells with widely variable DNA contents, including cells that lack macronuclei entirely. Protein expressed by the btu1-1 allele is dominant over wild-type protein expressed by the BTU2 locus. Normal macronuclear division is restored when the btu1-1 allele is inactivated by targeted disruption or expressed as a truncated protein. Immunofluorescence studies reveal elongated microtubular structures that surround macronuclei that fail to migrate to the cleavage furrows. In contrast, other cytoplasmic microtubule-dependent processes, such as cytokinesis, cortical patterning, and oral apparatus assembly, appear to be unaffected in the mutant. Micronuclear division is also perturbed in the K350M mutant, producing nuclei with elongated early-anaphase spindle configurations that persist well after the initiation of cytokinesis. The K350M mutation affects tubulin dynamics, as the macronuclear division defect is exacerbated by three treatments that promote microtubule polymerization: (i) elevated temperatures, (ii) sublethal concentrations of paclitaxel, and (iii) high concentrations of dimethyl sulfoxide. Inhibition of phosphatidylinositol 3-kinase (PI 3-kinase) with 3-methyladenine or wortmannin also induces amacronucleate cell formation in a btu1-1-dependent manner. Conversely, the myosin light chain kinase inhibitor ML-7 has no effect on nuclear division in the btu1-1 mutant strain. These findings provide new insights into microtubule dynamics and link the evolutionarily conserved PI 3-kinase signaling pathway to nuclear migration and/or division in Tetrahymena.

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Chronic fluoxetine treatment selectively uncouples raphe 5-HT(1A) receptors as measured by [(35)S]-GTP gamma S autoradiography.

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