Changes in ocular aquaporin expression following optic nerve crush.
OBJECTIVE
Changes in the expression of water channels (aquaporins; AQP) have been reported in several diseases. However, such changes and mechanisms remain to be evaluated for retinal injury after optic nerve crush (ONC). This study was designed to analyze changes in the expression of AQP4 (water selective channel) and AQP9 (water and lactate channel) following ONC in the rat.


METHODS
Rat retinal ganglion cells (RGCs) were retrogradely labeled by applying FluoroGold onto the left superior colliculus 1 week before ONC. Retinal injuries were induced by ONC unilaterally. Real-time PCR was used to measure changes in AQP4, AQP9, thy-1, Kir4.1 (K(+) channel), and beta-actin messages. Changes in AQP4, AQP9, Kir4.1, B cell lymphoma-x (bcl-xl), and glial fibrillary acidic protein (GFAP) expression were measured in total retinal extracts using western blotting.


RESULTS
The number of RGCs labeled retrogradely from the superior colliculus was 2,090+/-85 cells/mm(2) in rats without any treatment, which decreased to 1,091+/-78 (47% loss) and 497+/-87 cells/mm(2) (76% loss) on days 7 and 14, respectively. AQP4, Kir4.1, and thy-1 protein levels decreased at days 2, 7, and 14, which paralleled a similar reduction in mRNA levels, with the exception of Kir4.1 mRNA at day 2 showing an apparent upregulation. In contrast, AQP9 mRNA and protein levels showed opposite changes to those observed for the latter targets. Whereas AQP9 mRNA increased at days 2 and 14, protein levels decreased at both time points. AQP9 mRNA decreased at day 7, while protein levels increased. GFAP (a marker of astrogliosis) remained upregulated at days 2, 7, and 14, while bcl-xl (anti-apoptotic) decreased.


CONCLUSIONS
The reduced expression of AQP4 and Kir4.1 suggests dysfunctional ion coupling in retina following ONC and likely impaired retinal function. The sustained increase in GFAP indicates astrogliosis, while the decreased bcl-xl protein level suggests a commitment to cellular death, as clearly shown by the reduction in the RGC population and decreased thy-1 expression. Changes in AQP9 expression suggest a contribution of the channel to retinal ganglion cell death and response of distinct amacrine cells known to express AQP9 following traumatic injuries.
Dibas
Adnan
A
Department of Pharmacology & Neuroscience, University of North Texas Health Science Center at Fort Worth, Fort Worth, TX, USA. adibas@hsc.unt.edu
Oku
Hidehiro
H
Fukuhara
Masayuki
M
Kurimoto
Takuji
T
Ikeda
Tsunehiko
T
Patil
Rajkumar V
RV
Sharif
Najam A
NA
Yorio
Thomas
T
D000818:Animals; D051401:Aquaporin 4 / Q000235:genetics / Q000378:metabolism*; D020346:Aquaporins / Q000235:genetics / Q000378:metabolism*; D002452:Cell Count; D005123:Eye / Q000378:metabolism* / Q000473:pathology; D005786:Gene Expression Regulation; D005904:Glial Fibrillary Acidic Protein / Q000378:metabolism; D008297:Male; D009409:Nerve Crush; D009900:Optic Nerve / Q000378:metabolism* / Q000473:pathology / Q000601:surgery*; D024661:Potassium Channels, Inwardly Rectifying / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D012165:Retinal Ganglion Cells / Q000378:metabolism / Q000473:pathology; D018800:Thy-1 Antigens / Q000235:genetics / Q000378:metabolism; D051020:bcl-X Protein / Q000378:metabolism; D000098854:Kcnj10 Channel
C496032:Aqp4 protein, rat; C491637:Aqp9 protein, rat; D051401:Aquaporin 4; D020346:Aquaporins; D005904:Glial Fibrillary Acidic Protein; D000098854:Kcnj10 Channel; D024661:Potassium Channels, Inwardly Rectifying; D012333:RNA, Messenger; D018800:Thy-1 Antigens; D051020:bcl-X Protein