Pharmacological Characterization of a Serotonin Receptor (5-HT7) Stimulating cAMP Production in Human Corneal Epithelial Cells
To study the mRNA and pharmacology of a serotonin (5-HT) receptor positively coupled to adenylyl cyclase in normal, primary (P-CEPI), and immortalized human corneal epithelial cells (CEPI-17-CL4), by using numerous 5-HT agonists and antagonists. To determine and compare cloned human 5-HT7 receptor b...
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| Veröffentlicht in: | Investigative ophthalmology & visual science 2003-11, Vol.44 (11), p.4837-4844 |
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| creator | Crider, Julie Y Williams, Gary W Drace, Colene D Katoli, Parvaneh Senchyna, Michelle Sharif, Najam A |
| description | To study the mRNA and pharmacology of a serotonin (5-HT) receptor positively coupled to adenylyl cyclase in normal, primary (P-CEPI), and immortalized human corneal epithelial cells (CEPI-17-CL4), by using numerous 5-HT agonists and antagonists. To determine and compare cloned human 5-HT7 receptor binding affinities of compounds with their functional potency data.
RT-PCR was used to detect the presence of an mRNA for the human 5-HT7 receptor in CEPI-17-CL4 cells. Receptor-mediated production of cAMP in cultured cells was measured using an enzyme immunoassay. Compound binding affinities were determined using [3H]-lysergic acid diethylamide ([3H]-LSD) binding to cell membranes of human embryonic kidney (HEK-293) cells expressing the cloned human 5-HT7 receptor.
RT-PCR revealed the presence of a 5-HT7 receptor mRNA in CEPI-17-CL4 cells. Normal P-CEPI cells generated cAMP in response to 5-HT (-log EC50; pEC50=7.6), 5-carboxamidotryptamine (5-CT; pEC50=7.8), 5-methoxy-tryptamine (pEC50=7.0) and 5-methoxy-dimethyl-tryptamine (pEC50=5.7). In CEPI-17-CL4 cells, serotonergic agonists also stimulated cAMP production with different potencies (pEC50): 5-CT (7.4)>5-HT (6.5)> or =5-methoxy-tryptamine (6.1)>5-methoxy-dimethyl-tryptamine (5.4)> or =8-OH-DPAT (spiperone (7.4)> or =clozapine (7.2)=SB-258719 (7.2)>mianserin (6.9)>ketanserin (6.3). Antagonist pKi values in P-CEPI cells were methiothepin (8.7), spiperone (7.4) and SB-258719 (6.6). The rank order of affinity for displacement of [3H]-LSD from the cloned human 5-HT7 receptor was: methiothepin>ritanserin>mesulergine=clozapine> or =metergoline=5-HT>SB-258719> or =spiperone>mianserin> or =ketanserin. The functional agonist and antagonist potency data obtained from CEPI-17-CL4 cells correlated well with cloned human 5-HT7 receptor binding affinity data (r=0.69), with P-CEPI cell functional data (r=0.85), and with functional potency data in the literature for the cloned human 5-HT7 receptor (r=0.88).
These collective data support the presence of a pharmacologically defined, adenylyl cyclase-coupled 5-HT7 receptor in the CEPI-17-CL4 cells that may have relevance to physiological and/or pathologic functions of 5-HT7 receptors in the human cornea. |
| doi_str_mv | 10.1167/iovs.02-1292 |
| format | Article |
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RT-PCR was used to detect the presence of an mRNA for the human 5-HT7 receptor in CEPI-17-CL4 cells. Receptor-mediated production of cAMP in cultured cells was measured using an enzyme immunoassay. Compound binding affinities were determined using [3H]-lysergic acid diethylamide ([3H]-LSD) binding to cell membranes of human embryonic kidney (HEK-293) cells expressing the cloned human 5-HT7 receptor.
RT-PCR revealed the presence of a 5-HT7 receptor mRNA in CEPI-17-CL4 cells. Normal P-CEPI cells generated cAMP in response to 5-HT (-log EC50; pEC50=7.6), 5-carboxamidotryptamine (5-CT; pEC50=7.8), 5-methoxy-tryptamine (pEC50=7.0) and 5-methoxy-dimethyl-tryptamine (pEC50=5.7). In CEPI-17-CL4 cells, serotonergic agonists also stimulated cAMP production with different potencies (pEC50): 5-CT (7.4)>5-HT (6.5)> or =5-methoxy-tryptamine (6.1)>5-methoxy-dimethyl-tryptamine (5.4)> or =8-OH-DPAT (<5.0)=alpha-methyl-5-HT (<5.0). Various 5-HT receptor antagonists inhibited cAMP production induced by 5-CT in CEPI-17-CL4 cells with different potencies (pKi): methiothepin (8.5)>mesulergine (8.1)=metergoline (8.0)>spiperone (7.4)> or =clozapine (7.2)=SB-258719 (7.2)>mianserin (6.9)>ketanserin (6.3). Antagonist pKi values in P-CEPI cells were methiothepin (8.7), spiperone (7.4) and SB-258719 (6.6). The rank order of affinity for displacement of [3H]-LSD from the cloned human 5-HT7 receptor was: methiothepin>ritanserin>mesulergine=clozapine> or =metergoline=5-HT>SB-258719> or =spiperone>mianserin> or =ketanserin. The functional agonist and antagonist potency data obtained from CEPI-17-CL4 cells correlated well with cloned human 5-HT7 receptor binding affinity data (r=0.69), with P-CEPI cell functional data (r=0.85), and with functional potency data in the literature for the cloned human 5-HT7 receptor (r=0.88).
These collective data support the presence of a pharmacologically defined, adenylyl cyclase-coupled 5-HT7 receptor in the CEPI-17-CL4 cells that may have relevance to physiological and/or pathologic functions of 5-HT7 receptors in the human cornea.</description><identifier>ISSN: 0146-0404</identifier><identifier>ISSN: 1552-5783</identifier><identifier>EISSN: 1552-5783</identifier><identifier>DOI: 10.1167/iovs.02-1292</identifier><identifier>PMID: 14578406</identifier><identifier>CODEN: IOVSDA</identifier><language>eng</language><publisher>Rockville, MD: ARVO</publisher><subject>Adenylyl Cyclases - metabolism ; Adult ; Biological and medical sciences ; Cell Line ; Cell Membrane - metabolism ; Cyclic AMP - biosynthesis ; Epithelium, Corneal - drug effects ; Epithelium, Corneal - metabolism ; Eye and associated structures. Visual pathways and centers. Vision ; Fundamental and applied biological sciences. Psychology ; Humans ; Kidney - embryology ; Lysergic Acid Diethylamide - metabolism ; Middle Aged ; Receptors, Serotonin - genetics ; Receptors, Serotonin - metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; RNA, Messenger - metabolism ; Serotonin Antagonists - pharmacology ; Serotonin Receptor Agonists - pharmacology ; Vertebrates: nervous system and sense organs</subject><ispartof>Investigative ophthalmology & visual science, 2003-11, Vol.44 (11), p.4837-4844</ispartof><rights>2004 INIST-CNRS</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c349t-fe3adf4e09f52079d2f6860fa9b5143662bd2433d6061ae98e0e3803b2e391f13</citedby></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>314,780,784,27924,27925</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=15249009$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/14578406$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Crider, Julie Y</creatorcontrib><creatorcontrib>Williams, Gary W</creatorcontrib><creatorcontrib>Drace, Colene D</creatorcontrib><creatorcontrib>Katoli, Parvaneh</creatorcontrib><creatorcontrib>Senchyna, Michelle</creatorcontrib><creatorcontrib>Sharif, Najam A</creatorcontrib><title>Pharmacological Characterization of a Serotonin Receptor (5-HT7) Stimulating cAMP Production in Human Corneal Epithelial Cells</title><title>Investigative ophthalmology & visual science</title><addtitle>Invest Ophthalmol Vis Sci</addtitle><description>To study the mRNA and pharmacology of a serotonin (5-HT) receptor positively coupled to adenylyl cyclase in normal, primary (P-CEPI), and immortalized human corneal epithelial cells (CEPI-17-CL4), by using numerous 5-HT agonists and antagonists. To determine and compare cloned human 5-HT7 receptor binding affinities of compounds with their functional potency data.
RT-PCR was used to detect the presence of an mRNA for the human 5-HT7 receptor in CEPI-17-CL4 cells. Receptor-mediated production of cAMP in cultured cells was measured using an enzyme immunoassay. Compound binding affinities were determined using [3H]-lysergic acid diethylamide ([3H]-LSD) binding to cell membranes of human embryonic kidney (HEK-293) cells expressing the cloned human 5-HT7 receptor.
RT-PCR revealed the presence of a 5-HT7 receptor mRNA in CEPI-17-CL4 cells. Normal P-CEPI cells generated cAMP in response to 5-HT (-log EC50; pEC50=7.6), 5-carboxamidotryptamine (5-CT; pEC50=7.8), 5-methoxy-tryptamine (pEC50=7.0) and 5-methoxy-dimethyl-tryptamine (pEC50=5.7). In CEPI-17-CL4 cells, serotonergic agonists also stimulated cAMP production with different potencies (pEC50): 5-CT (7.4)>5-HT (6.5)> or =5-methoxy-tryptamine (6.1)>5-methoxy-dimethyl-tryptamine (5.4)> or =8-OH-DPAT (<5.0)=alpha-methyl-5-HT (<5.0). Various 5-HT receptor antagonists inhibited cAMP production induced by 5-CT in CEPI-17-CL4 cells with different potencies (pKi): methiothepin (8.5)>mesulergine (8.1)=metergoline (8.0)>spiperone (7.4)> or =clozapine (7.2)=SB-258719 (7.2)>mianserin (6.9)>ketanserin (6.3). Antagonist pKi values in P-CEPI cells were methiothepin (8.7), spiperone (7.4) and SB-258719 (6.6). The rank order of affinity for displacement of [3H]-LSD from the cloned human 5-HT7 receptor was: methiothepin>ritanserin>mesulergine=clozapine> or =metergoline=5-HT>SB-258719> or =spiperone>mianserin> or =ketanserin. The functional agonist and antagonist potency data obtained from CEPI-17-CL4 cells correlated well with cloned human 5-HT7 receptor binding affinity data (r=0.69), with P-CEPI cell functional data (r=0.85), and with functional potency data in the literature for the cloned human 5-HT7 receptor (r=0.88).
These collective data support the presence of a pharmacologically defined, adenylyl cyclase-coupled 5-HT7 receptor in the CEPI-17-CL4 cells that may have relevance to physiological and/or pathologic functions of 5-HT7 receptors in the human cornea.</description><subject>Adenylyl Cyclases - metabolism</subject><subject>Adult</subject><subject>Biological and medical sciences</subject><subject>Cell Line</subject><subject>Cell Membrane - metabolism</subject><subject>Cyclic AMP - biosynthesis</subject><subject>Epithelium, Corneal - drug effects</subject><subject>Epithelium, Corneal - metabolism</subject><subject>Eye and associated structures. Visual pathways and centers. Vision</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>Humans</subject><subject>Kidney - embryology</subject><subject>Lysergic Acid Diethylamide - metabolism</subject><subject>Middle Aged</subject><subject>Receptors, Serotonin - genetics</subject><subject>Receptors, Serotonin - metabolism</subject><subject>Reverse Transcriptase Polymerase Chain Reaction</subject><subject>RNA, Messenger - metabolism</subject><subject>Serotonin Antagonists - pharmacology</subject><subject>Serotonin Receptor Agonists - pharmacology</subject><subject>Vertebrates: nervous system and sense organs</subject><issn>0146-0404</issn><issn>1552-5783</issn><issn>1552-5783</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2003</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNpF0M1v0zAYBnALgVgZ3DgjX_iSyHj9ESc-TtVGkYao2DhbrmO3Rk7c2QkVO_C349BKO1mWfn4e60HoNYELQkTz2cff-QJoRaikT9CC1DWt6qZlT9ECCBcVcOBn6EXOvwAoIRSeozPCi-AgFujveqdTr00MceuNDnhZ7tqMNvkHPfo44Oiwxrc2xTEOfsA_rLH7MSb8oa5Wd81HfDv6fgrFDltsLr-t8TrFbjL_3xa_mno94GVMgy3pV3s_7mzwc5ENIb9Ez5wO2b46nefo5_XV3XJV3Xz_8nV5eVMZxuVYOct057gF6WoKjeyoE60Ap-WmJpwJQTcd5Yx1AgTRVrYWLGuBbahlkjjCztG7Y-4-xfvJ5lH1PpvyAz3YOGXVlPVkLWSBn47QpJhzsk7tk-91-qMIqHlvNe-tgKp578LfnHKnTW-7R3wauIC3J6BzmdclPRifH11NuQSYe98f3c5vdwefrMq9DqHEEnU4HDgv5Yq3rGH_ANctloM</recordid><startdate>20031101</startdate><enddate>20031101</enddate><creator>Crider, Julie Y</creator><creator>Williams, Gary W</creator><creator>Drace, Colene D</creator><creator>Katoli, Parvaneh</creator><creator>Senchyna, Michelle</creator><creator>Sharif, Najam A</creator><general>ARVO</general><general>Association for Research in Vision and Ophtalmology</general><scope>IQODW</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7X8</scope></search><sort><creationdate>20031101</creationdate><title>Pharmacological Characterization of a Serotonin Receptor (5-HT7) Stimulating cAMP Production in Human Corneal Epithelial Cells</title><author>Crider, Julie Y ; Williams, Gary W ; Drace, Colene D ; Katoli, Parvaneh ; Senchyna, Michelle ; Sharif, Najam A</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c349t-fe3adf4e09f52079d2f6860fa9b5143662bd2433d6061ae98e0e3803b2e391f13</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2003</creationdate><topic>Adenylyl Cyclases - metabolism</topic><topic>Adult</topic><topic>Biological and medical sciences</topic><topic>Cell Line</topic><topic>Cell Membrane - metabolism</topic><topic>Cyclic AMP - biosynthesis</topic><topic>Epithelium, Corneal - drug effects</topic><topic>Epithelium, Corneal - metabolism</topic><topic>Eye and associated structures. Visual pathways and centers. Vision</topic><topic>Fundamental and applied biological sciences. Psychology</topic><topic>Humans</topic><topic>Kidney - embryology</topic><topic>Lysergic Acid Diethylamide - metabolism</topic><topic>Middle Aged</topic><topic>Receptors, Serotonin - genetics</topic><topic>Receptors, Serotonin - metabolism</topic><topic>Reverse Transcriptase Polymerase Chain Reaction</topic><topic>RNA, Messenger - metabolism</topic><topic>Serotonin Antagonists - pharmacology</topic><topic>Serotonin Receptor Agonists - pharmacology</topic><topic>Vertebrates: nervous system and sense organs</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Crider, Julie Y</creatorcontrib><creatorcontrib>Williams, Gary W</creatorcontrib><creatorcontrib>Drace, Colene D</creatorcontrib><creatorcontrib>Katoli, Parvaneh</creatorcontrib><creatorcontrib>Senchyna, Michelle</creatorcontrib><creatorcontrib>Sharif, Najam A</creatorcontrib><collection>Pascal-Francis</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>MEDLINE - Academic</collection><jtitle>Investigative ophthalmology & visual science</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Crider, Julie Y</au><au>Williams, Gary W</au><au>Drace, Colene D</au><au>Katoli, Parvaneh</au><au>Senchyna, Michelle</au><au>Sharif, Najam A</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Pharmacological Characterization of a Serotonin Receptor (5-HT7) Stimulating cAMP Production in Human Corneal Epithelial Cells</atitle><jtitle>Investigative ophthalmology & visual science</jtitle><addtitle>Invest Ophthalmol Vis Sci</addtitle><date>2003-11-01</date><risdate>2003</risdate><volume>44</volume><issue>11</issue><spage>4837</spage><epage>4844</epage><pages>4837-4844</pages><issn>0146-0404</issn><issn>1552-5783</issn><eissn>1552-5783</eissn><coden>IOVSDA</coden><abstract>To study the mRNA and pharmacology of a serotonin (5-HT) receptor positively coupled to adenylyl cyclase in normal, primary (P-CEPI), and immortalized human corneal epithelial cells (CEPI-17-CL4), by using numerous 5-HT agonists and antagonists. To determine and compare cloned human 5-HT7 receptor binding affinities of compounds with their functional potency data.
RT-PCR was used to detect the presence of an mRNA for the human 5-HT7 receptor in CEPI-17-CL4 cells. Receptor-mediated production of cAMP in cultured cells was measured using an enzyme immunoassay. Compound binding affinities were determined using [3H]-lysergic acid diethylamide ([3H]-LSD) binding to cell membranes of human embryonic kidney (HEK-293) cells expressing the cloned human 5-HT7 receptor.
RT-PCR revealed the presence of a 5-HT7 receptor mRNA in CEPI-17-CL4 cells. Normal P-CEPI cells generated cAMP in response to 5-HT (-log EC50; pEC50=7.6), 5-carboxamidotryptamine (5-CT; pEC50=7.8), 5-methoxy-tryptamine (pEC50=7.0) and 5-methoxy-dimethyl-tryptamine (pEC50=5.7). In CEPI-17-CL4 cells, serotonergic agonists also stimulated cAMP production with different potencies (pEC50): 5-CT (7.4)>5-HT (6.5)> or =5-methoxy-tryptamine (6.1)>5-methoxy-dimethyl-tryptamine (5.4)> or =8-OH-DPAT (<5.0)=alpha-methyl-5-HT (<5.0). Various 5-HT receptor antagonists inhibited cAMP production induced by 5-CT in CEPI-17-CL4 cells with different potencies (pKi): methiothepin (8.5)>mesulergine (8.1)=metergoline (8.0)>spiperone (7.4)> or =clozapine (7.2)=SB-258719 (7.2)>mianserin (6.9)>ketanserin (6.3). Antagonist pKi values in P-CEPI cells were methiothepin (8.7), spiperone (7.4) and SB-258719 (6.6). The rank order of affinity for displacement of [3H]-LSD from the cloned human 5-HT7 receptor was: methiothepin>ritanserin>mesulergine=clozapine> or =metergoline=5-HT>SB-258719> or =spiperone>mianserin> or =ketanserin. The functional agonist and antagonist potency data obtained from CEPI-17-CL4 cells correlated well with cloned human 5-HT7 receptor binding affinity data (r=0.69), with P-CEPI cell functional data (r=0.85), and with functional potency data in the literature for the cloned human 5-HT7 receptor (r=0.88).
These collective data support the presence of a pharmacologically defined, adenylyl cyclase-coupled 5-HT7 receptor in the CEPI-17-CL4 cells that may have relevance to physiological and/or pathologic functions of 5-HT7 receptors in the human cornea.</abstract><cop>Rockville, MD</cop><pub>ARVO</pub><pmid>14578406</pmid><doi>10.1167/iovs.02-1292</doi><tpages>8</tpages></addata></record> |
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| subjects | Adenylyl Cyclases - metabolism Adult Biological and medical sciences Cell Line Cell Membrane - metabolism Cyclic AMP - biosynthesis Epithelium, Corneal - drug effects Epithelium, Corneal - metabolism Eye and associated structures. Visual pathways and centers. Vision Fundamental and applied biological sciences. Psychology Humans Kidney - embryology Lysergic Acid Diethylamide - metabolism Middle Aged Receptors, Serotonin - genetics Receptors, Serotonin - metabolism Reverse Transcriptase Polymerase Chain Reaction RNA, Messenger - metabolism Serotonin Antagonists - pharmacology Serotonin Receptor Agonists - pharmacology Vertebrates: nervous system and sense organs |
| title | Pharmacological Characterization of a Serotonin Receptor (5-HT7) Stimulating cAMP Production in Human Corneal Epithelial Cells |
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