Characterization of the Mel1c melatoninergic receptor in platypus (<i>Ornithorhynchus anatinus</i>) - Fig 5

<p><b>Molecular pharmacology of the Mel1c receptors from platypus (A) and clawed frog (B).</b> The ligand was 2-[¹²⁵I]iodomelatonin. Independent experiments were performed at least twice using different batches of membranes from stably transfected CHO cells and each point was obtained in triplicate. SD6, N-[2-(5-methoxy-1H-indol-3-yl)ethyl]iodoacetamide; 2IMLT, 2-iodomelatonin; 6-Cl- Chloro MLT, 6-chloromelatonin; Luzindole, N-acetyl-2-Br-MLT; 2-bromomelatonin, benzyltryptamine; 4P-P-DOT, N-[(2S,4S)-4-phenyl-1,2,3,4-tetrahydronaphthalen-2-yl]propanamide; Agomelatin^® (S20098), N-(2-(7-methoxynaphthalen-1-yl)ethyl)acetamide; Ramelteon^® (FLN68), (S)-N-(2-(1,6,7,8-tetrahydro-2H-indeno-(5,4)furan-8-yl)ethyl)propionamide; D600, methoxyverapamil; 5HT, 5-hydroxytryptamine; S20928, (N-[2-(1-naphthyl)ethyl] cyclobutanecarboxamide); S21278, N-[2-(6-methoxybenzimidazol-1-yl)ethyl]acetamide; S22153, N-[2-(5-ethylbenzothiophen-3-yl)ethyl]acetamide; S27128-1, N-[2-(2-iodo-5-methoxy-6-nitro-1H-indol-3-yl)ethyl]acetamide; S73893, N-[3-methoxy-2-(7-methoxy-1-naphthyl)propyl]acetamide; S75436, 2-fluoro-N-[3-hydroxy-2-(7-methoxy-1-naphthyl) propyl]acetamide; S77834S27128, N-[(8-[2-(2-iodo-5-methoxy-10,11-dihydro-5H-dibenzo[a,d][<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0191904#pone.0191904.ref007" target="_blank">7</a>] annulen-106-nitro-1H-indol-3-yl)methylethyl]acetamide; S77840, 1-[(8-methoxy-10,11-dihydro-5H-dibenzo[a,d][<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0191904#pone.0191904.ref007" target="_blank">7</a>]annulen-101H-indol-3-yl)methyl]urea ethyl]iodoacetamide; SD1881, N-[2-(6-iodo-5-methoxy-1H-indol-3-yl)ethyl]acetamideiodomelatonin; SD1882, N-[2-(4-iodo-5-methoxy-1H-indol-3-yl)ethyl]acetamide; SD1918, N-[7-iodomelatonin; Div 880, 2-(7-iodo-5-methoxy-1H-indol-3-yl)ethyl]acetamide2-[(2-iodo-4,5-dimethoxyphenyl)methyl]-4,5-dimethoxy phenyl; S70254, 2-iodo-N-2-[5-methoxy-2-(naphthalen-1-yl)-1H-pyrrolo[3,2-b]pyridine-3-yl])acetamide. Concentration isotherms were obtained using 10 concentrations of each product from 10⁻¹³ to 10⁻⁴ M.</p

Comparison of Mel1c sequences in platypus, xenopus and chicken to the human GPR50.

<p>Comparison of Mel1c sequences in platypus, xenopus and chicken to the human GPR50.</p

Characterization of the Mel1c melatoninergic receptor in platypus (<i>Ornithorhynchus anatinus</i>) - Fig 7

<p><b>Comparison of the correlations between the molecular pharmacology of platypus Mel1c (A) or clawed frog Mel1c (B) and human melatonin receptors MT</b>_<b>1</b><b>and MT</b>_<b>2</b>. The correlation in red is between hMT₁ and platypus Mel1c; in blue, between hMT₂ and Platypus Mel1c (upper panel). The correlation in red is between hMT₁ and clawed frog Mel1c; and in green between hMT₂ and clawed frog Mel1c (lower panel.) See <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0191904#pone.0191904.g005" target="_blank">Fig 5</a> for data.</p

Molecular pharmacology of Mel1c receptors in platypus and xenopus compared to the human MT₁ and MT₂ melatonin receptors.

<p>Molecular pharmacology of Mel1c receptors in platypus and xenopus compared to the human MT₁ and MT₂ melatonin receptors.</p

Expression of Mel1c receptors in COS7 cells.

<p>Subcellular localization of HA epitope-tagged Mel1c receptors. Immunofluorescence studies were performed with transfected COS7cells. COS7 cells and COS7 expressing 3HA-platypus Mel1C or 3HA-X. laevis Mel1C or 3HA-G. gallus Mel1C receptors were probed with mouse monoclonal anti-HA antibody directed against the N-terminal epitope tag present on recombinant receptor. Experiments were carried out with paraformaldehyde-fixed and non-permeabilized cells. The fluorescence images were obtained by using Alexa 488-conjugated goat anti- mouse IgG secondary antibody. COS7 cell nuclei were stained with 4′, 6′-diamidino-2-phenylindole. Native COS7 cells were used as a negative control.</p

Saturation binding experiments for 2-[125I]-iodomelatonin.

<p>Membranes from COS7 cells transfected with platypus Mel1c (A), clawed frog Mel1c (B), and chicken Mel1c (C) were used to measure the binding at Mel1c receptors. Red line represents specific binding, black line represents total binding and dotted line represents non-specific binding.</p

Comparison of the molecular pharmacology of platypus, xenopus and chicken melatonin Mel1c receptors.

<p>Comparison of the molecular pharmacology of platypus, xenopus and chicken melatonin Mel1c receptors.</p

Therian mammals used for phylogenetic analysis.

<p>Therian mammals used for phylogenetic analysis.</p

Primer sequences used in this study.

<p>Primer sequences used in this study.</p

Correlation between the data obtained from platypus Mel1c and those obtained with clawed frog Mel1c.

<p>Data were obtained in a binding assay with 2-[¹²⁵I]-iodomelatonin as the radioligand. See <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0191904#pone.0191904.g005" target="_blank">Fig 5</a>.</p