Minimaal-invasieve techniek voor injectie in Rat Optic Nerve
Summary
Direct injection into the rat optic nerve is useful for regenerative research. We demonstrate a minimally-invasive technique for direct injection into a rat optic nerve that does not involve opening the skull. Using this method, surgical complications are minimized and recovery is more rapid.
Abstract
The rat optic nerve is a useful model for stem cell regeneration research. Direct injection into the rat optic nerve allows delivery into the central nervous system in a minimally-invasive surgery without bone removal. This technique describes an approach to visualization and direct injection of the optic nerve following minor fascial dissection from the orbital ridge, using a conjunctival traction suture to gently pull the eye down and out. Representative examples of an injected optic nerve show successful injection of dyed beads.
Introduction
De oogzenuw biedt een ideale locatie voor het centrale zenuwstelsel (CNS) regeneratief onderzoek met inbegrip van oogheelkundige aandoeningen, zoals optische neuritis, glaucoom en trauma. Injecties van verschillende stamcellen zijn ofwel aangetoonde werkzaamheid of belofte getoond bij het vervangen van verloren myeline, waardoor axonale tellen en / of voorkomen van degeneratieve ziekten. 1,2
De menselijke oogzenuw bevat ongeveer 1.200.000 parallel axonen op weg van de retina naar de chiasm met een diameter van ongeveer 3,0-3,5 mm. 3 Voor menselijke ziekten te modelleren in het laboratorium, is de rat vaak toegepast. De volwassen rat oogzenuw bevat ongeveer 100.000 axonen binnen een diameter van ongeveer 0,5 mm. 4 een van de belangrijkste beperkingen van CNS regeneratieve activiteit is direct been. Complicaties en chirurgische risico's voor het dier hoger wanneer de schedel of wervels verwijderd. Net als de voordelen vanminimaal invasieve benaderingen in de ruggengraat, 5 direct oogzenuw injecties zonder de schedel aanbod verlaagde complicaties en een sneller herstel.
Deze techniek is gebruikt in eerdere studies. 6 In dit manuscript en bijbehorende video, demonstreren we een minimaal invasieve procedure stamcellen injecteren in de ratten optische zenuw.
Protocol
Representative Results
Discussion
Direct injection into the optic nerve of stem cells or other products intended to facilitate regeneration provides a convenient model compared to other means of injections into the CNS. This technique takes less time, requires less total anesthesia, avoids drilling or removing skull or bone tissue, reduces complications rates and allows for more rapid recovery following surgery.
The most critical steps in this protocol include: 1. Adequate hemostasis in the surgical field to allow clear visua…
Divulgations
The authors have nothing to disclose.
Acknowledgements
This study was supported by NeuralStem, Inc., and Johns Hopkins Project RESTORE.
Materials
| Name of Material/ Equipment | Company | Catalog Number | Comments/Description |
| Lewis rat | Charles River | 4 | Any rat strain will work. |
| Anesthesia machine | Surgivet | CDS9000 | CDS 9000 Small Animal Anesthesia Machine – Pole Mount |
| Infusion pump | Stoelting | 53129 | |
| Dissection microscope | National Optical | 409-411-1105 | |
| Fiber-optic light source | Fisher Scientific | 12-562-21 | |
| Dissection and Stereotaxic Instrument | Stoelting | 51400 | |
| Pipette Puller | Kopf | 750 | |
| Pipettes | World Precision Instruments | 18150-6 | |
| Disposable scalpel blades | Harvard Apparatus | 810-15-021 | |
| Iridectomy scissors | Electron Microscopy Sciences | Uniband LA-4XF |
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