Expression of adenylate cyclase subtypes II and IV in the human outflow pathway.
OBJECTIVE
It has been demonstrated that low doses of pilocarpine and other muscarinics substantially increase outflow facility in the isolated human outflow system devoid of ciliary muscle. These cholinergic-induced facility responses were thought possibly to be due to elevation of cAMP as a result of the presence of adenylate cyclases II (AC-II) and IV (AC-IV). Therefore, whether these isoforms are present in outflow tissues was examined.


METHODS
Human anterior segments were perfused with carbachol (10(-9)-10(-5) M), and outflow facility and cAMP levels in the perfusate were measured simultaneously. Isolated trabecular meshwork (TM) were incubated with carbachol (10(-7) M), and the subsequent changes in cAMP were measured by radioimmunoassay. AC-II and AC-IV were characterized in ocular tissue with reverse transcription-polymerase chain reaction and in situ hybridization.


RESULTS
Outflow facility increased, in a dose-dependent manner, by 10%, 16%, and 27% in response to 10(-9), 10(-7), and 10(-5) M carbachol, respectively. Similarly, cAMP increased by 9%, 70%, and 210% in response to 10(-9), 10(-7), and 10(-5) M carbachol, respectively. In addition, cAMP levels significantly increased by 39% in isolated TM strips incubated with 10(-7) M carbachol. AC-II was detected in most normal tissue examined, but not in any cultured cell lines or any glaucomatous tissue. AC-IV was also widely expressed in most normal tissues, faintly detected in some glaucoma tissue, but not detected in most cultured cells.


CONCLUSIONS
The presence of AC-II and AC-IV in outflow tissues supports the hypothesis that cholinergics may indeed exert an effect on outflow facility, mediated by cAMP, which is independent of muscle contraction.
Zhang
X
X
Boston University School of Medicine, Department of Ophthalmology, Massachusetts 02118, USA.
Wang
N
N
Schroeder
A
A
Erickson
K A
KA
D000262:Adenylyl Cyclases / Q000096:biosynthesis / Q000235:genetics*; D000368:Aged; D000369:Aged, 80 and over; D002217:Carbachol / Q000494:pharmacology; D002478:Cells, Cultured; D018679:Cholinergic Agonists / Q000494:pharmacology; D002924:Ciliary Body / Q000187:drug effects / Q000201:enzymology*; D003315:Cornea / Q000187:drug effects / Q000201:enzymology; D000242:Cyclic AMP / Q000378:metabolism; D017931:DNA Primers / Q000737:chemistry; D015870:Gene Expression; D005901:Glaucoma / Q000201:enzymology* / Q000473:pathology; D006801:Humans; D017403:In Situ Hybridization; D012333:RNA, Messenger / Q000378:metabolism; D011863:Radioimmunoassay; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012590:Sclera / Q000187:drug effects / Q000201:enzymology; D014129:Trabecular Meshwork / Q000187:drug effects / Q000201:enzymology*
D018679:Cholinergic Agonists; D017931:DNA Primers; D012333:RNA, Messenger; D002217:Carbachol; D000242:Cyclic AMP; D000262:Adenylyl Cyclases; C100622:adenylyl cyclase 2; C406262:adenylyl cyclase 4