Loading...
Activation of stretch-activated channels and maxi-K+ channels by membrane stress of human lamina cribrosa cells.
Irnaten, Mustapha Barry, Richard C Quill, Barry Clark, Abbot F Harvey, Brian J P O'Brien, Colm J
Irnaten, Mustapha
Barry, Richard C
Quill, Barry
Clark, Abbot F
Harvey, Brian J P
O'Brien, Colm J
Citations
Altmetric:
Advisors
Editors
Other Contributors
Date
2009-01
Date Submitted
Keywords
Other Subjects
Subject Mesh
Biological Markers
Calcium
Cell Culture Techniques
Cell Membrane
Cell Size
DNA Primers
Gadolinium
Humans
Hypotonic Solutions
Ion Channels
Large-Conductance Calcium-Activated Potassium Channel alpha Subunits
Large-Conductance Calcium-Activated Potassium Channels
Male
Microscopy, Confocal
Optic Disk
Patch-Clamp Techniques
Peptides
RNA, Messenger
Reverse Transcriptase Polymerase Chain Reaction
Scorpion Venoms
Stress, Physiological
Thapsigargin
Calcium
Cell Culture Techniques
Cell Membrane
Cell Size
DNA Primers
Gadolinium
Humans
Hypotonic Solutions
Ion Channels
Large-Conductance Calcium-Activated Potassium Channel alpha Subunits
Large-Conductance Calcium-Activated Potassium Channels
Male
Microscopy, Confocal
Optic Disk
Patch-Clamp Techniques
Peptides
RNA, Messenger
Reverse Transcriptase Polymerase Chain Reaction
Scorpion Venoms
Stress, Physiological
Thapsigargin
Planned Date
Start Date
Collaborators
Principal Investigators
Files
Loading...
z7g00109000194.pdf
Adobe PDF, 388.81 KB
Alternative Titles
Publisher
Abstract
The lamina cribrosa (LC) region of the optic nerve head is considered the primary site of damage in glaucomatous optic neuropathy. Resident LC cells have a profibrotic potential when exposed to cyclical stretch. However, the mechanosensitive mechanisms of these cells remain unknown. Here the authors investigated the effects of membrane stretch on cell volume change and ion channel activity and examined the associated changes in intracellular calcium ([Ca(2+)](i)).
The authors used primary LC cells obtained from normal human donor eyes. Confocal microscopy was used to investigate the effect of hypotonic cell membrane stretch on cell volume changes. Whole-cell patch-clamp and calcium imaging techniques were used to investigate the effect of hypotonicity on ion channel(s) activity and [Ca(2+)](i) changes, respectively. RT-PCR was used to examine for the maxi-K(+) signature in LC cells.
In this study, LC cells showed significant volume changes in response to hypotonic cell swelling. The authors characterized a large conductance K(+) channel (maxi-K(+)) in LC cells and demonstrated its increased activity during cell membrane hypotonic stretch. RT-PCR revealed the presence of maxi-K(+) signature in LC cells. The authors showed the [Ca(2+)](i) and maxi-K(+) channels to be dependent on extracellular Ca(2+) and inhibited by gadolinium, which blocks stretch-activated channels (SACs). Pretreatment with thapsigargin, which blocks the release of Ca(2+) from endoplasmic reticulum stores, showed no significant difference in [Ca(2+)](i) concentration on hypotonic swelling.
The results show that hypotonic stress of human LC cells activates SAC and Ca(2+)-dependent maxi-K(+) channels and that the increase in [Ca(2+)](i) during cell swelling was predominantly from extracellular sources (or intracellular stores other than the endoplasmic reticulum). These findings improve the understanding of how LC cells respond to cell membrane stretch. Further experiments in this area may reveal future targets for novel therapeutic intervention in the management of glaucoma.
The authors used primary LC cells obtained from normal human donor eyes. Confocal microscopy was used to investigate the effect of hypotonic cell membrane stretch on cell volume changes. Whole-cell patch-clamp and calcium imaging techniques were used to investigate the effect of hypotonicity on ion channel(s) activity and [Ca(2+)](i) changes, respectively. RT-PCR was used to examine for the maxi-K(+) signature in LC cells.
In this study, LC cells showed significant volume changes in response to hypotonic cell swelling. The authors characterized a large conductance K(+) channel (maxi-K(+)) in LC cells and demonstrated its increased activity during cell membrane hypotonic stretch. RT-PCR revealed the presence of maxi-K(+) signature in LC cells. The authors showed the [Ca(2+)](i) and maxi-K(+) channels to be dependent on extracellular Ca(2+) and inhibited by gadolinium, which blocks stretch-activated channels (SACs). Pretreatment with thapsigargin, which blocks the release of Ca(2+) from endoplasmic reticulum stores, showed no significant difference in [Ca(2+)](i) concentration on hypotonic swelling.
The results show that hypotonic stress of human LC cells activates SAC and Ca(2+)-dependent maxi-K(+) channels and that the increase in [Ca(2+)](i) during cell swelling was predominantly from extracellular sources (or intracellular stores other than the endoplasmic reticulum). These findings improve the understanding of how LC cells respond to cell membrane stretch. Further experiments in this area may reveal future targets for novel therapeutic intervention in the management of glaucoma.
Language
en
ISSN
1552-5783
eISSN
ISBN
DOI
10.1167/iovs.08-1937
PMID
18775862