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Biomedical subjects

J T Laitinen

Publications and source records attributed to J T Laitinen.

At least 19 recordsLinked to original sources
PubMed2003

Effects of mobile phone radiation on UV-induced skin tumourigenesis in ornithine decarboxylase transgenic and non-transgenic mice.

PURPOSE: The effects of low-level radiofrequency radiation (RFR) on ultraviolet (UV)-induced skin tumorigenesis were evaluated in ornithine decarboxylase (ODC) and non-transgenic mice. MATERIALS AND METHODS: Transgenic female mice over-expressing the human ODC gene and their non-transgenic littermates (20 animals in the cage control group, and 45-49 animals in the other groups) were exposed for 52 weeks to UV radiation or a combination of UV radiation and pulsed RFR. The UV dose was 240 Jm(-2) (1.2 x human minimum erythemal dose) delivered three times a week. One group of animals was exposed to Digital Advanced Mobile Phone System (DAMPS)-type RFR, the other group to Global System for Mobile (GSM)-type RFR at a nominal average specific absorption rate of 0.5 W kg(-1), 1.5 h day(-1), for 5 days a week. The skin was carefully palpated weekly for macroscopic tumours. Histopathological analyses of all skin lesions and of a specified dorsal skin area were performed on all animals. RESULTS: UV exposure resulted in development of macroscopic skin tumours in 11.5 and 36.8% of non-transgenic and transgenic animals, respectively. The RFR exposures did not give a statistically significant effect on the development of skin tumours in either transgenic or non-transgenic animals, or in combined analysis, but tumour development appeared slightly accelerated especially in non-transgenic animals. No effects of RFR exposures were found on excretion of 6-hydroxymelatonin sulphate into urine or on polyamine levels in dorsal skin. CONCLUSION: RFR exposures did not significantly enhance skin tumourigenesis. However, the slightly accelerated tumour development may warrant further evaluation.

Animals↗
PubMed2001

Effects of topical anandamide-transport inhibitors, AM404 and olvanil, on intraocular pressure in normotensive rabbits.

PURPOSE: To evaluate the effects of topically applied anandamide transport inhibitors, AM404 and olvanil, on the intraocular pressure (IOP) of normotensive rabbits. To determine if the ocular hypotension induced by topical anandamide (AEA) can be potentiated by co-administered AM404. METHODS: Test compounds, in either hydroxypropyl-beta-cyclodextrin (HP-beta-CD) or propylene glycol, were administered unilaterally onto rabbit eyes. To determine if AM404 affects the IOP-profile of AEA, AM404 was administered ocularly 15 minutes before topical AEA. Phenylmethylsulfonyl fluoride (PMSF) (24 mg/kg, s.c.) was given 30 min before AEA to prevent its catabolism. IOPs of the treated and untreated eyes were measured. The cannabinoid agonist activities of AM404 and olvanil were studied by using [35S]GTPyS autoradiography. RESULTS: Topical AM404 (62.5 micirog), in HP-beta-CD vehicle, decreased IOP significantly in treated eyes. AM404 (62.5 microg) induced a significant IOP increase without subsequent decrease when given in propylene glycol vehicle. Olvanil (312.5 microg) caused a significant IOP reduction without provoking an initial hypertensive phase. These compounds did not significantly affect the IOP of untreated eyes. Co-administered AM404 (125 microg in HP-beta-CD) had no significant effect on the IOP profile of AEA (62.5 microg). CONCLUSIONS: Ocular administration of AM404 or olvanil decreased IOP in rabbits, although AM404 can provoke an initial ocular hypertension and did not potentiate the IOP responses induced by exogenous AEA.

Administration, Topical↗
PubMed2001

Despite substantial degradation, 2-arachidonoylglycerol is a potent full efficacy agonist mediating CB(1) receptor-dependent G-protein activation in rat cerebellar membranes.

1. Two endocannabinoids, arachidonoyl ethanolamide (AEA) and 2-arachidonoylglycerol (2-AG) bind and activate G-protein-coupled cannabinoid receptors, but limited data exist on their relative ability to activate G-proteins. 2. Here we assess agonist potency and efficacy of various cannabinoids, including 2-AG, HU-310 (2-arachidonoyl glyceryl ether, a third putative endocannabinoid), HU-313 (another ether analogue of 2-AG), AEA, R-methanandamide (an enzymatically stable analogue of AEA), and CP-55,940 at rat brain CB(1) receptors using agonist-stimulated [(35)S]-GTPgammaS binding to cerebellar membranes and whole brain sections. Degradation of endocannabinoids under experimental conditions was monitored by HPLC. 3. To enhance efficacy differences, agonist dose-response curves were generated using increasing GDP concentrations. At 10(-6) M GDP, all compounds, except HU-313, produced full agonists responses approximately 2.5 fold over basal. The superior efficacy of 2-AG over all other compounds became evident by increasing GDP (10(-5) and 10(-4) M). 4. In membrane incubations, 2-AG was degraded by 85% whereas AEA and HU-310 were stable. Pretreatment of membranes with phenylmethylsulphonyl fluoride inhibited 2-AG degradation, resulting in 2 fold increase in agonist potency. Such pretreatment had no effect on AEA potency. 5. Responses in brain sections were otherwise consistent with membrane binding data, but 2-AG evoked only a weak signal in brain sections, apparently due to more extensive degradation. 6. These data establish that even under conditions of substantial degradation, 2-AG is a full efficacy agonist, clearly more potent than AEA, in mediating CB(1) receptor-dependent G-protein activity in native membranes.

Animals↗
PubMed2001

[(35)S]GTPgammaS autoradiography reveals a wide distribution of G(i/o)-linked ADP receptors in the nervous system: close similarities with the platelet P2Y(ADP) receptor.

No G(i)-linked P2Y receptors have been cloned to date but the presence of such receptors is thought to be restricted to platelets and certain clonal cell lines. Using the functional approach of [(35)S]guanosine 5'-[gamma-thio]-triphosphate autoradiography, we uncovered the widespread presence of such receptors in the CNS. Under conditions in which the prominent signal due to tonic adenosine receptor activity is masked, ADP and ATP stimulated G-protein activity in multiple grey and white matter regions. Localization in the grey matter suggests inhibitory auto-/heteroreceptor function. In the white matter, activated G proteins appeared as 'hot spots' (presumed oligodendrocyte progenitors) with scattered distribution along the main fibre tracts. Responses to ATP were diminished under conditions that inhibited degradation, suggesting that prior conversion to ADP explained agonist action. Uracil nucleotides were ineffective but 2-methylthio-ADP activated G proteins approximately 500-fold more potently than ADP, although both were similarly degraded. Throughout the brain, ADP-dependent G-protein activity was reversed by 2-hexylthio-AdoOC(O)Asp(2), a non-phosphate ATP analogue, whereas selective P2Y(1) receptor antagonists proved ineffective. A similar receptor was also disclosed from the adrenal medulla. These data witness a hitherto unrecognized abundance of G(i/o)-linked ADP receptors in the nervous system. Biochemical and pharmacological behaviour suggests striking similarities to the elusive platelet P2Y(ADP) receptor.

Adenine Nucleotides↗
PubMed2000

2,3,7,8-Tetrachlorodibenzo-p-dioxin alters melatonin metabolism in fish hepatocytes.

Pineal hormone melatonin is an important regulator of endocrine and circadian rhythms in vertebrates. Since liver is assumed to be the major organ in the metabolism of this indole hormone, we investigated the effect of the known Ah-receptor agonist, 2,3,7, 8-tetrachlorodibenzo-p-dioxin (TCDD) on melatonin metabolism in fish hepatocytes as well as the in vitro effect of melatonin on trout hepatic microsomal cytochrome P4501A (CYP1A) catalyst. Primary cell cultures of rainbow trout hepatocytes were exposed to [3H]melatonin (1 nM to 1 microM) alone and in combination with TCDD (50 pM) at 15 degrees C for 24 or 48 h. Analysis of melatonin and its metabolites in the culture medium and hepatocytes by HPLC revealed that about 96% of the added [3H]melatonin was metabolised after 24 h in both control and TCDD treated cultures. 3H-radioactivity was found mainly in the culture medium and less than 5% of the total 3H-radioactivity retained inside hepatocytes. Of the HPLC separated metabolites, one coeluted with 6-hydroxymelatonin and one unknown metabolite eluted after 6-hydroxymelatonin. In addition, two other metabolites were more water-soluble than 6-hydroxymelatonin and were considered to be conjugated products. Treatment of the hepatocytes with TCDD increased the amount of the major oxidated product, 6-hydroxymelatonin, about 2.5-fold after 24 h and 1.2-fold after 48 h exposure, respectively when compared with the control cultures. Whereas the amount of the unknown metabolite eluting after 6-hydroxymelatonin decreased about 1.3-fold after 24 h and 1.2-fold after 48 h exposure, respectively. Melatonin alone did not affect P4501A associated EROD-activity or CYP1AmRNA levels in the primary hepatocyte cultures. TCDD-treatment increased EROD-activity 3 to 5-fold and respective CYP1AmRNA content 6 to 14-fold, when compared with the control or melatonin-treated cultures. Furthermore, melatonin competitively inhibited EROD-activity in liver microsomes with a Ki value of 62.06+/-3.78 microM. The results show that TCDD alters metabolic degradation of melatonin in hepatocytes and suggest that P4501A may be an important P450 isoenzyme involved in oxidative metabolism of melatonin in fish liver.

Animals↗
PubMed2000

Overexpression of heme oxygenase in neuronal cells, the possible interaction with Tau.

Increased expression of heme oxygenase-1 (HO-1) is a common feature in a number of neurodegenerative diseases. Interestingly, the spatial distribution of HO-1 expression in diseased brain is essentially identical to that of pathological expression of tau. In this study, we explored the relationship between HO-1 and tau, using neuroblastoma cells stably transfected with sense and antisense HO-1 constructs as well as with the vector alone. In transfected cells overexpressing HO-1, the activity of heme oxygenase was increased, and conversely, the level of tau protein was dramatically decreased when compared with antisense HO-1 or CEP transfected cells. The suppression of tau protein expression was almost completely reversed by zinc-deuteroporphyrin, a specific inhibitor of heme oxygenase activity. The activated forms of ERKs (extracellular signal-regulated kinases) were also decreased in cells overexpressing HO-1 although no changes in the expression of total ERK-1/2 proteins were observed. These data are in agreement with the finding that the expression of tau is regulated through signal cascades including the ERKs, whose activities are modulated by oxidative stresses. The expression of tau and HO-1 may be regulated by oxidative stresses in a coordinated manner and play a pivotal role in the cytoprotection of neuronal cells.

Alzheimer Disease↗
PubMed2000

Nocturnal 6-hydroxymelatonin sulfate excretion in female workers exposed to magnetic fields.

The objective of this study was to determine whether daytime occupational exposure to extremely low frequency magnetic fields (MFs) suppresses nocturnal melatonin production. Sixty female volunteers were recruited. Thirty-nine worked in a garment factory, and 21 office workers served as a reference group. Exposure assessment was based on the type of sewing machine used and MF measurements around each type of machine. Eye-level MF flux density was used to classify the operators to higher (>1 microT) and lower (0.3-1 microT) exposure categories. A third group of factory workers had diverse MF exposures from other sources. The reference group had average exposure of about 0.15 microT. Urine samples were collected on Friday and Monday for three consecutive weeks. Melatonin production was assessed as urinary 6-hydroxymelatonin sulfate (6-OHMS) excretion. The ratio of Friday morning/Monday morning 6-OHMS was used to test the hypothesis that melatonin production is suppressed after 4 days of occupational MF exposure with significant recovery during the weekend. Possible chronic suppression of melatonin production was evaluated by studying exposure-related differences in the Friday values by multivariate regression analysis. The Monday/Friday ratios were close to 1.0, suggesting that there is no increase in melatonin production over the weekend. The average 6-OHMS excretion on Friday was lower among the factory workers than in the reference group, but no monotonous dose-response was observed. Multivariate regression analysis identified MF exposure, smoking, and age as significant explanatory variables associated with decreased 6-OHMS excretion.

Adult↗
PubMed1999

Synthesis and in vitro pharmacology of a series of new chiral histamine H3-receptor ligands: 2-(R and S)-Amino-3-(1H-imidazol-4(5)-yl)propyl ether derivatives.

To investigate stereospecificity and the mechanism of activation of the histamine H3-receptor, a series of 2-(R and S)-amino-3-(1H-imidazol-4(5)-yl)propyl ether derivatives were synthesized. In these compounds, the structures of the well-known antagonist iodoproxyfan and the full agonists R- or S-(alpha)-methylhistamine were combined in one molecule. The obtained "hybrid" molecules were tested for H3-receptor affinity on rat cerebral cortex. Some selected compounds were further screened for H3-receptor functional activity with GTPgamma[35S] autoradiography studies using rat brain tissue sections. The affinity of all the synthesized compounds (-log Ki = 5.9-7.9) was lower than that found for iodoproxyfan or two of its analogues; however, the compounds showed stereospecificity. The S-configuration of the series of 2-amino-3-(1H-imidazol-4(5)-yl)propyl ether derivatives, which resembles the stereochemistry of R-(alpha)-methylhistamine, was more favorable. Incorporation of an amino group in the propyl chain of iodoproxyfan and analogues did not alter the antagonistic behavior for compounds with an aromatic side chain. However, when also the aromatic moiety was replaced by a cyclohexyl group, the compounds behaved as agonists. This indicates that an interaction between the side chain amino group and the H3-receptor protein is involved in H3-receptor activation. The 2-(S)-amino-3-(1H-imidazol-4(5)-yl)propyl cyclohexylmethyl ether (23) has H3-receptor agonistic properties with high affinity for the histamine H3-receptor (-log Ki = 7.9 +/- 0.2) and might serve as a useful tool for further studies concerning drug design and receptor-ligand interactions.

Animals↗
PubMed1999

The effect of photoperiod on diel rhythms in serum melatonin, cortisol, glucose, and electrolytes in the common dentex, Dentex dentex.

Diel rhythms in serum concentrations of melatonin, cortisol, glucose, sodium, chloride, and potassium were studied in the common dentex, Dentex dentex, under different photoperiods (DD, 8L:16D, 12L:12D, 16L:8D). Photoperiod affected both the diel rhythms and the absolute values of the estimated blood components. Regardless of the photoperiod, melatonin titers were elevated during the scotophase (384.3 +/- 13.9 pg/ml) compared with a mean baseline level of 54.4 +/- 2.7 pg/ml during the photophase. Serum melatonin concentrations reflected the prevailing photoperiod and constantly elevated melatonin levels with no diel rhythmicity were evident in fish held in the DD protocol. A circadian-like pattern in serum cortisol was observed in fish that were kept at the DD and 8L:16D protocols with cortisol peak at 18:00 h in the night. Fish exposed to the 16L:8D regime showed highest cortisol levels at 10:00 h, while no rhythmicity was evident under the 12L:12D protocol. A phase shift of 4 h between the peaks of cortisol and glucose was evident in fish exposed to the DD, 8L:16D, and 12L:12D regimes. Diel patterns of changes in serum Na+ and Cl- were observed only in the fish held in the DD protocol. Serum K+ values were lowest during the first part of the scotophase under all regimes, except the 16L:8D where no diel rhythmicity was detected. During the photophase, cortisol was positively correlated with glucose, Na+, and Cl- and negatively with K+. During the scotophase, melatonin was positively correlated with glucose and electrolytes. Results indicated that cortisol may be responsible for the observed rhythmicity of glucose and that melatonin may play a role in glucose and ion regulation in common dentex.

Animals↗
PubMed1999

Effects of continuous opioid receptor blockade on alcohol intake and up-regulation of opioid receptor subtype signalling in a genetic model of high alcohol drinking.

Effects of a continuous naloxone infusion via osmotic pumps on alcohol drinking and opioid receptor density and function in the high-drinking AA (Alko, Alcohol) rats were examined. AA rats were trained to drink 10% (v/v) ethanol in a 1-h limited access procedure and implanted with subcutaneous osmotic pumps delivering either saline, a low dose (0.3 mg/kg per hour), or a high dose (3.0 mg/kg per hour) of naloxone for 7 days. The pumps were then removed and alcohol, food and water intakes were measured for another 4 days. Compared with saline, both naloxone doses significantly suppressed 1-h alcohol intake during the 7-day infusion. The suppression was smaller than that by a bolus injection of the same daily dose 15 min before the session, although a complete blockade of morphine-induced antinociception was achieved even with the smaller naloxone infusion. Significant decreases were also seen in daily food and water intake during the first days, but they quickly returned to their previous baselines. After pump removal, rats of both naloxone-treated groups rapidly increased their alcohol drinking and reached the pretreatment baseline, while their food and water intakes significantly surpassed their baselines. Naloxone infusion at 3.0 mg/kg per hour for 7 days significantly decreased 24-h alcohol drinking without affecting alcohol preference. Twenty-four hours after pump removal, autoradiography with [3H]DAMGO, [3H]DPDPE and [3H]U-69,543 revealed an up-regulation of mu-, delta- and kappa-opioid receptor binding sites in many brain areas of these animals. This receptor up-regulation was functional, because receptor coupling to G-protein activation was enhanced by agonist ligands, as revealed by [35S]GTPgammaS autoradiography. A good correlation existed between ligand binding densities and G-protein activation for mu- and kappa-receptors in control and naloxone-treated brain sections. Furthermore, morphine-induced analgesia in a hot-plate test showed a leftward shift in the morphine dose-response curve after naloxone treatment. These results suggest that the usefulness of a chronic opioid antagonist dosing regime could be limited by nonspecific effects of the antagonist on ingestive behaviour, an up-regulation of opioid receptors with high antagonist doses, and the resulting supersensitivity to opioid agonists after the discontinuation of the treatment.

Alcohol Drinking↗
PubMed1999

Selective detection of adenosine A1 receptor-dependent G-protein activity in basal and stimulated conditions of rat brain [35S]guanosine 5'-(gamma-thio)triphosphate autoradiography.

[35S]Guanosine 5'-(gamma-thio)triphosphate autoradiography is a novel technique to detect receptor-dependent activation of G-proteins in brain tissue sections. While an increasing number of reports using this approach are beginning to appear, little effort has been directed to the identification of factors responsible for the heterogeneously distributed [35S]guanosine 5'-(gamma-thio)triphosphate signal in basal conditions. The present study demonstrates that endogenously formed adenosine generates a widespread and prominent adenosine A1 receptor-dependent signal in basal conditions using this technique. Treatment of rat brain tissue sections with the A1-selective antagonist 8-cyclopentyl-1,3-dipropylxanthine dose-dependently (EC50 < 10 nM) suppressed basal [35S]guanosine 5'-(gamma-thio)triphosphate binding in a region-specific manner, an effect fully mimicked by the adenosine-depleting enzyme adenosine deaminase, and less so by the A1 antagonist cirsimarin and by caffeine. That adenosine was continuously formed during the incubation is supported by the constant requirements of adenosine deaminase in order to suppress basal radioligand binding and further by the fact that low micromolar concentrations of adenine nucleotides evoked only adenosine-mimicking and fully 8-cyclopentyl-1,3-dipropylxanthine-sensitive binding responses. In the presence of adenosine deaminase, all responses to adenine nucleotides were abolished, indicating that prior conversion to adenosine was required. Upon stimulation, this technique selectively detected A1 receptor-activated G-proteins, as the non-selective agonists adenosine and 2-chloroadenosine and the A1-selective agonist N6-p-sulfophenyladenosine all evoked only 8-cyclopentyl-1,3-dipropylxanthine-sensitive responses in identical gray matter areas, and also in several white matter areas such as the corpus callosum, anterior commissure, optic tract and cerebellar white matter. Dose-response studies revealed region-specific differences in the magnitude of A1 receptor-stimulated G-protein activation, with the highest response (nine-fold over basal) detectable in the hippocampus. No response to the A2A-selective agonist 2-[(2-aminoethylamino)carbonylethylphenylethylamino]-5'-N-et hylcarboxamidoadenosine or the A3-selective agonist 2-chloro-N6-(3-iodobenzyl)-adenosine-5'-N-methyluronamide was detected in any region. These data reveal that a significant amount of noise inherent to [35S]guanosine 5'-(gamma-thio)triphosphate autoradiography can be eliminated by removal of the adenosine signal, a step likely facilitating detection of responses to other receptors. Furthermore, the data establish [35S]guanosine 5-(gamma-thio)triphosphate autoradiography as a novel and selective approach to directly assess A1 receptor-G-protein coupling in anatomically defined regions of the central nervous system.

Adenosine Deaminase↗
PubMed1998

Mutagenesis of human Mel1a melatonin receptor expressed in yeast reveals domains important for receptor function.

A yeast functional colorimetric assay was employed to test the effects of site-directed point mutations on the function of the human Mel1a melatonin receptor. Seven mutants were created in transmembrane domains III, V, and VII of the receptor to test the rhodopsin-based model of melatonin recognition. Two mutants in transmembrane domains III and VI were created to investigate the mechanisms of G protein activation in the melatonin receptor. Mutations in transmembrane domain V either potentiated agonist efficiencies (H195A) or totally abolished all responses to tested compounds (V192T+H195A). Mutation N124A in the conserved NRY motif in the end of transmembrane domain III seriously impaired receptor activation. Several mutants were found to have decreased ability to activate functional responses, reflecting the importance of these residues for receptor function. These data also suggest that activation of the receptor involves interaction of the 5-methoxy group of melatonin with the conserved histidine H195 in transmembrane domain V.

Binding Sites↗
PubMed1998

Guanosine 5'-(gamma-[35S]thio)triphosphate autoradiography allows selective detection of histamine H3 receptor-dependent G protein activation in rat brain tissue sections.

Histamine elicits its biological effects via three distinct G protein-coupled receptors, termed H1, H2, and H3. We have used guanosine 5'-(gamma-[35S]thio)triphosphate (GTPgamma[35S]) autoradiography to localize histamine receptor-dependent G protein activation in rat brain tissue sections. Initial studies revealed that in basal conditions, adenosine was present in tissue sections in sufficient concentrations to generate an adenosine A1 receptor-dependent GTPgamma[35S] signal in several brain regions. All further incubations therefore contained 8-cyclopentyl-1,3-dipropylxanthine (10 microM), a selective A1 receptor antagonist. Histamine elicited dose-dependent increments in GTPgamma[35S] binding to discrete anatomical structures, most notably the caudate putamen, cerebral cortex, and substantia nigra. The overall anatomical pattern of the histamine-evoked binding response closely reflects the known distribution of H3 binding sites and was faithfully mimicked by N(alpha)-methylhistamine, (R)-alpha-methylhistamine, and immepip, three H3-selective agonists. In all regions examined, the GTPgamma[35S] signal was reversed with thioperamide and clobenpropit, two potent H3-selective antagonists, whereas mepyramine, a specific H1 antagonist, and cimetidine, a prototypic H2 antagonist, proved ineffective. These data indicate that in rat brain tissue sections, GTPgamma[35S] autoradiography selectively detects H3 receptor-dependent signaling in response to histamine stimulation. As the existing evidence suggests that GTPgamma[35S] autoradiography preferentially reveals responses to G(i/o)-coupled receptors, our data indicate that most, if not all, central H3 binding sites represent functional receptors coupling to G(i/o), the inhibitory class of G proteins. Besides allowing more detailed studies on H3 receptor signaling within anatomically restricted regions of the CNS, GTPgamma[35S] autoradiography offers a novel approach for functional in vitro screening of H3 ligands.

Adenosine↗
PubMed1998

Protective properties of tin- and manganese-centered porphyrins against hydrogen peroxide-mediated injury in rat astroglial cells.

Tin-mesoporphyrin (tin-mp), a potent inhibitor of heme oxygenase, and manganese (III) tetrakis(1-methyl-4-pyridyl)porphyrin (MnTMPyP), a potent superoxide dismutase mimetic, reduced H2O2 toxicity in cultures of transformed rat astroglial cells if added 30 min before, or at the same time as, H2O2. Reduced toxicity was not observed if treatment was delayed for 60 min, the time by which H2O2 was essentially eliminated from cultures. Coadministration of tin-mp and MnTMPyP did not increase protection over either compound administered individually. Tin-mp, but not MnTMPyP, was stable in culture. MnCl2 was not protective, suggesting that protection by MnTMPyP was not dependent on manganous ion, a by-product of MnTMPyP breakdown. Protection by tin-mp and MnTMPyP was not associated with metalloporphyrin-mediated induction of heme oxygenase-1 or with changes in heme oxygenase-2 on western blots. Whereas protective concentrations of tin-mp did not have superoxide dismutase-mimetic properties in vitro, protective concentrations of MnTMPyP partially inhibited heme oxygenase. The data support the hypothesis that heme oxygenase inhibition is protective against acute oxidative injury.

Animals↗
PubMed1998

Melatonin receptor genes.

Melatonin is produced rhythmically by the pineal gland and the retina with increased synthesis during darkness. Pineal melatonin serves as the 'chemical expression of darkness' conveying information on the ambient light-dark cycle into rhythmic bodily functions. On-going debate on modes and sites of action ranges from views of melatonin affecting each and every cell ('cure-all') to those of melatonin having restricted actions through specific high-affinity receptors. The present review deals with the latter view. The use of 2-[125I]-iodomelatonin has allowed the exact localization and characterization of high-affinity melatonin receptors that signal through the G(i/o) class of G proteins. Molecular cloning of melatonin receptor genes has confirmed that most, if not all, high-affinity melatonin-binding sites represent the G-protein-coupled melatonin receptors. Based on sequence dissimilarities, melatonin receptors are classified into three subtypes, Mel1a, Mel1b and Mel1c. A distribution wider than originally thought of melatonin receptors in the human brain and peripheral sites has brought these receptors into focus of several drug companies, promising exciting times for research on melatonin and new therapeutic possibilities.

Amino Acid Sequence↗
PubMed1998

Diurnal rhythm of melatonin in bovine milk: pharmacokinetics of exogenous melatonin in lactating cows and goats.

We investigated whether the melatonin levels in bovine milk exhibit a similar daily rhythm as serum levels. In 4 Ayrshire cows at the beginning of the lactation period in May the nocturnal rise in milk melatonin was moderate (from 7 +/- 2 pg/ml at noon to 15 +/- 1 pg/ml at night; mean +/- SEM) and did not correlate well with the melatonin level in serum (from 7 +/- 2 pg/ml to 27 +/- 7 pg/ml, respectively). On the other hand, 6 cows in a later phase of lactation, studied in February, showed a clear long-lasting nocturnal melatonin increase both in serum (from 9 +/- 1 pg/ml at noon to 26 +/- 3 pg/ml at night) and in milk (from 12 +/- 5 pg/ml to 26 +/- 7 pg/ml, respectively). Melatonin kinetics during lactation was studied in more detail in 4 Ayrshire cows and 4 dairy goats by giving an intravenous bolus injection of melatonin. A 3-compartment model with melatonin elimination from the central compartment was used to describe the data. The values (mean +/- SD) for the cows and the goats were: elimination half-life 27 +/- 4 min and 27 +/- 1 min, mean residence time 24 +/- 4 min and 18 +/- 4 min, steady state distribution volume 1.0 +/- 0.3 l/kg and 0.6 +/- 0.1 1/kg (p < 0.05), and plasma clearance 0.044 +/- 0.004 l/kg/min and 0.035 +/- 0.011 l/kg/min, respectively. Following injection, the melatonin concentration in milk increased rapidly and exceeded the corresponding serum value 15-30 min later, remaining thereafter above the serum level. Our results suggest that milk melatonin levels reflect blood concentrations of melatonin with a short delay.

Animal Husbandry↗
PubMed1997

[Gamma-35S]GTP autoradiography allows region-specific detection of muscarinic receptor-dependent G-protein activation in the chick optic tectum.

A recently introduced technique of [gamma-35S]GTP autoradiography was used to localize and characterize muscarinic receptor-dependent activation of G-proteins in tissue sections of the chick optic tectum, a brain region with relatively high expression of G-protein-coupled receptors for the neurotransmitter acetylcholine. Within the highly stratified tectal structure, the bulk of muscarinic receptor-mediated [gamma-35S]GTP signal was localized to the stratum griseum et fibrosum superficiale with considerably lower binding responses in other tectal layers. Quantitative comparison of [gamma-35S]GTP binding responses in tectal sections and membranes revealed a close match between the two tissue preparations for the response elicited by the cholinergic agonist carbachol, its dose-dependent reversal with the non-selective muscarinic antagonist atropine, its approximately 100-fold sensitivity towards blockade with M1-type (pirenzepine) over M2-type (gallamine) muscarinic antagonists, as well as absolute requirement for micromolar concentrations of GDP (EC50 approximately 10 microM) for the receptor-mediated [gamma-35S]GTP response. The pharmacological profile is consistent with that of cm4, a recently cloned chicken homolog of the mammalian m4 muscarinic acetylcholine receptor. Moreover, the strict GDP-dependence of the binding response suggest activation of Gi/o, the inhibitory class of G-proteins. These data provide the first functional characterization of the chick tectal muscarinic receptors. A close match between [gamma-35S]GTP responses in membranes and tissue sections strongly suggest that [gamma-35S]GTP autoradiography offers great potential for studies on G-protein-mediated signaling, with particular use within anatomically restricted regions that are not readily approached with more conventional techniques. It is anticipated that [gamma-35S]GTP autoradiography should greatly facilitate studies on signaling capacity of an individual receptor subtype whilst in its native cellular environment.

Animals↗