Expression and functional characterization of a melatonin‐sensitive receptor in <i>Xenopus</i> oocytes

Fischer, Bernhard; Muβhoff, U.; Fauteck, Jan-Dirk; Madeja, Michael; Wittkowski, W.; Speckmann, Erwin‐Josef

doi:10.1016/0014-5793(96)00090-7

Cited by 5 publications

(9 citation statements)

References 26 publications

(38 reference statements)

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“…2), whereas native oocytes or water-injected oocytes did not respond to melatonin (data not shown). The time course of expression of melatonin receptors in oocytes following mRNA injection was similar to the one observed for melatonin receptors after injection of mRNA from the whole rat brain and from hippocampal tissue [24][25][26]. Maximum reponses occurred between 3 and 5 days after mRNA injection.…”

Section: Resultssupporting

confidence: 68%

Gene expression and functional characterization of melatonin receptors in the spinal cord of the rat: implications for pain modulation

Zahn¹,

Lansmann

Berger

et al. 2003

Journal of Pineal Research

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Recently, a species-dependent distribution of melatonin binding sites have been found in lamina I-V and lamina X of the spinal cord. In order to learn more about the function of spinal melatonin receptors, we investigated (i) the gene expression for melatonin receptor subtypes in lumbar and thoracal spinal cord tissue by means of the reverse-transcriptase polymerase chain reaction (RT-PCR) technique, and (ii) the electrophysiological and pharmacological properties of melatonin receptors heterologously expressed in Xenopus oocytes after injection of spinal cord mRNA by means of the voltage clamp technique. Because ample evidence indicates an antinociceptive effect of melatonin, (iii) the role of spinal melatonin receptors for maintaining mechanical and thermal hyperalgesia was studied in a rat model for postoperative pain. The RT-PCR data revealed that transcripts for MT1 and MT2 melatonin receptors are present in the dorsal and ventral horn of lumbar and thoracal spinal cord tissue. Injection of mRNA from lumbar spinal cord tissue into Xenopus oocytes led to the functional reconstitution of melatonin receptors which activate calcium-dependent chloride inward currents. Melatonin responses were abolished by simultaneous administration of the antagonists, 2-phenylmelatonin and luzindole and were unaffected by the MT2 antagonist 4-phenyl-2-propionamidotetralin. Intrathecal administration of different melatonin doses (10-100 nmol) did not inhibit mechanical or thermal hyperalgesia. However, intrathecal application of a low dose of morphine together with melatonin caused a brief antinociceptive effect suggesting an enhanced morphine analgesia by melatonin. In conclusion, the present study demonstrated for the first time the presence of transcripts of MT1 and MT2 receptors located in the dorsal and ventral horn of the spinal cord. Furthermore, spinal melatonin enhanced the antinociceptive effect of morphine indicating that melatonin acts as a neuromodulator in the spinal cord.

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Section: Resultssupporting

confidence: 68%

Gene expression and functional characterization of melatonin receptors in the spinal cord of the rat: implications for pain modulation

Zahn¹,

Lansmann

Berger

et al. 2003

Journal of Pineal Research

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“…The administration of melatonin to the mRNA-injected oocytes evoked oscillatory inward currents ( Fig.3A; n ϭ 61), whereas native oocytes or water-injected oocytes did not respond to either melatonin or other neurotransmitters (data not shown; see Fischer et al, 1996). The time course of expression of melatonin receptors in oocytes following mRNA injection was similar to that observed for melatonin receptors after injection of mRNA from whole rat brain (Fischer et al, 1996;Blumenau et al, 2001). Maximum responses occurred between 3-5 days after mRNA injection.…”

Section: Xenopus Oocytesmentioning

confidence: 82%

“…Injection of mRNA into the oocytes led to the functional expression of various ligand-operated ion channels sensitive to the neurotransmitters ␥-aminobutyric acid, dopamine, serotonin, and glutamate with its agonists kainate, quisqualate, and N-methyl-D-aspartate, and AMPA (data not shown; see Mußhoff et al, 1994). The administration of melatonin to the mRNA-injected oocytes evoked oscillatory inward currents ( Fig.3A; n ϭ 61), whereas native oocytes or water-injected oocytes did not respond to either melatonin or other neurotransmitters (data not shown; see Fischer et al, 1996). The time course of expression of melatonin receptors in oocytes following mRNA injection was similar to that observed for melatonin receptors after injection of mRNA from whole rat brain (Fischer et al, 1996;Blumenau et al, 2001).…”

Section: Xenopus Oocytesmentioning

confidence: 82%

“…This melatonin-sensitive receptor was determined to be exogenous, since native oocytes or waterinjected oocytes were not responsive to melatonin. We previously reported the functional expression of a melatonin-sensitive receptor in Xenopus oocytes after injection of rat brain mRNA (Fischer et al, 1996;Blumenau et al, 2001). This receptor activates via the phospholipase C signaling pathway calcium-dependent chloride channels in the oocyte membrane, resulting in oscillatory chloride inward currents.…”

Section: Discussionmentioning

confidence: 99%

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Melatonin receptors in rat hippocampus: molecular and functional investigations

et al. 2002

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Since binding sites for melatonin have been found in the hippocampus of several mammals, it has been suggested that the pineal hormone melatonin is able to modulate neuronal functions of hippocampal cells. In order to get more insight into the role of melatonin for the functions of hippocampal cells, the following experiments were performed: male rats, maintained under a 12/12-h light-dark cycle, were sacrificed by decapitation at zeitgeber times (h) ZT2, ZT8, and ZT15 (ZT0 = lights on); for experiment 1, gene expression for melatonin receptors was detected in the hippocampus and in hippocampal subfields by means of the RT-PCR technique; for experiment 2, electrophysiological and pharmacological properties of melatonin receptors heterologously expressed in Xenopus oocytes after injection of mRNA from the hippocampus were analyzed by means of voltage clamp technique; and for experiment 3, effects of melatonin on the spontaneous firing rate of action potentials in the CA1 regions of hippocampal slices were analyzed by means of extracellular recordings. The RT-PCR data revealed that transcripts for both the MT1 and MT2 melatonin receptors are present in the dentate gyrus, CA3, and CA1 regions, and the subiculum of the hippocampus. Injection of mRNA from rat hippocampus into the Xenopus oocytes led to the functional reconstitution of melatonin-sensitive receptors, which activates calcium-dependent chloride inward currents. The melatonin responses were abolished by simultaneous administration of the antagonists 2-phenylmelatonin and luzindole, and were unaffected by the MT2 antagonist 4-phenyl-2-propionamidotetralin. Bath-applied melatonin (1 micromol/l) enhances the firing rate of neurons in the CA1 region. The effect was small in experiments performed at ZT8 (<2 times the initial level) and large in experiments performed at ZT15 (>6 times). The changes of neuronal firing rate induced by melatonin were completely suppressed with simultaneous administration of the melatonin receptor antagonist luzindole (10 micromol/l). The results indicate that melatonin may play an important role in modulating neuronal excitability in the hippocampus.

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“…Xenopus oocytes have been a useful system for expressing exogenous membrane receptors and ion channels (Sigel, 1990). Using this expression system, it has been demonstrated that melatonin receptors, when expressed from rat brain mRNA, can positively couple to the phospholipase C-mediated signaling pathway in the oocyte (Fischer et al, 1996). The increased phospholipase C activity leads to elevated levels of inositol 1,4,5-triphosphate (IP 3 ) and to the release of calcium from IP 3 -sensitive intracellular stores.…”

Section: Introductionmentioning

confidence: 99%

Expression and functional characterization of the mt₁ melatonin receptor from rat brain in Xenopus oocytes: evidence for coupling to the phosphoinositol pathway

Blumenau¹,

Berger²,

Fauteck³

et al. 2001

Journal of Pineal Research

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Melatonin-sensitive receptors were expressed in Xenopus laevis oocytes following an injection of mRNA from rat brain. The administration of 0.1-100 micromol/L melatonin to voltage-clamped oocytes activates calcium-dependent chloride currents via a pertussis toxin-sensitive G protein and the phosphoinositol pathway. To determine which melatonin receptor type (mt1, MT2, MT3) is functionally expressed in the Xenopus oocytes, we used (i) agonists and antagonists of different receptor types to characterize the pharmacological profile of the expressed receptors and (ii) a strategy of inhibiting melatonin receptor function by antisense oligonucleotides. During pharmacological screening administration of the agonists 2-iodomelatonin and 2-iodo-N-butanoyl-5-methoxytryptamine (IbMT) to the oocytes resulted in oscillatory membrane currents, whereas the administration of the MT3 agonist 5-methoxycarbonylamino-N-acetyltryptamine (GR135,531) exerted no detectable membrane currents. The melatonin response was abolished by a preceding administration of the antagonists 2-phenylmelatonin and luzindole but was unaffected by the MT3 antagonist prazosin and the MT2 antagonist 4-phenyl-2-propionamidotetralin (4-P-PDOT). In the antisense experiments, in the control group the melatonin response occurred in 45 of 54 mRNA-injected oocytes (83%). Co-injection of the antisense oligonucleotide, corresponding to the mt1 receptor mRNA, caused a marked and significant reduction in the expression level (13%; P < 0.001). In conclusion, the results demonstrate that injection of mRNA from rat brain in Xenopus oocytes induced the expression of the mt1 receptor which is coupled to the phosphoinositol pathway.

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Expression and functional characterization of a melatonin‐sensitive receptor in Xenopus oocytes

Cited by 5 publications

References 26 publications

Gene expression and functional characterization of melatonin receptors in the spinal cord of the rat: implications for pain modulation

Gene expression and functional characterization of melatonin receptors in the spinal cord of the rat: implications for pain modulation

Melatonin receptors in rat hippocampus: molecular and functional investigations

Expression and functional characterization of the mt₁ melatonin receptor from rat brain in Xenopus oocytes: evidence for coupling to the phosphoinositol pathway

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Expression and functional characterization of a melatonin‐sensitive receptor in Xenopus oocytes

Cited by 5 publications

References 26 publications

Gene expression and functional characterization of melatonin receptors in the spinal cord of the rat: implications for pain modulation

Gene expression and functional characterization of melatonin receptors in the spinal cord of the rat: implications for pain modulation

Melatonin receptors in rat hippocampus: molecular and functional investigations

Expression and functional characterization of the mt1 melatonin receptor from rat brain in Xenopus oocytes: evidence for coupling to the phosphoinositol pathway

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Expression and functional characterization of the mt₁ melatonin receptor from rat brain in Xenopus oocytes: evidence for coupling to the phosphoinositol pathway