Hippocampale Insulin Mikroinjektion og<em> In vivo</em> Mikrodialyse Under Spatial Memory Test
Summary
Modulation af hippocampally-afhængig rumlig arbejdshukommelse ved direkte intrahippocampal mikroinjektion, ledsaget og efterfulgt af<em> In vivo</em> Mikrodialyse for metabolitter i bevidste, opfører dyr.
Abstract
Glucosemetabolisme er en nyttig markør for lokal neural aktivitet, danner grundlag for metoder såsom 2-deoxyglucose og funktionel magnetisk resonans billeddannelse. Dog er brug af sådanne metoder i dyremodeller kræver anæstesi og dermed begge ændrer hjernen tilstand og forhindrer adfærdsmæssige foranstaltninger. En alternativ fremgangsmåde er anvendelsen af in vivo mikrodialyse at tage kontinuerlig måling af hjernen ekstracellulære væske koncentrationer af glucose, lactat og beslægtede metabolitter i vågen, uhæmmede dyr. Denne teknik er især nyttig, når den kombineres med opgaver, der skal basere sig på specifikke hjerneregioner og / eller akut farmakologisk manipulation, for eksempel, viser hippocampale målinger under en rumlig arbejdshukommelse opgave (spontan vekslen) et fald i ekstracellulær glucose og stigning i lactat, der er tyder på øget glykolyse 1,2, og intrahippocampal insulin administration både forbedrer hukommelsen og øger hippocampus glycolradiokemiske analyser 3. Stoffer såsom insulin kan leveres til hippocampus via samme mikrodialysesonde anvendes til at måle metabolitter. Brugen af spontan vekslen som et mål for hippocampus funktion er designet til at undgå enhver forvirre fra stressende motivatorer (f.eks fodchok), tilbageholdenhed, eller belønninger (fx fødevarer), som alle kan ændre både opgaveløsning og stofskifte, denne opgave giver også en måling af motorisk aktivitet, der tillader kontrol for ikke-specifikke virkninger af behandlingen. Kombineret, disse metoder tillader direkte måling af de neurokemiske og metaboliske variabler regulerer adfærd.
Protocol
Representative Results
Discussion
Alle opløsninger anvendt i mikrodialyse skal filtreres umiddelbart før brug under anvendelse af et 0,2 um filter. Efter indføring af sonden ind i føringen kanyle, observere at bekræfte, at strømningen er uhindret og prøveudtagning finder sted. Hvis flow stopper efter indsættelse, den mest sandsynlige årsag er skader på sonden membran forårsaget af utilstrækkelig pleje på indsættelse, og en frisk sonde skal erstattes.
Som bemærket ovenfor, en væsentlig fordel for disse fremgan…
Disclosures
The authors have nothing to disclose.
Acknowledgements
Dette arbejde blev støttet af NIH / NIDDK (DK077106 til ECM).
Materials
The majority of reagents are standard laboratory grade and can be obtained from a supplier of choice. Similarly, equipment such as syringe pumps and tubing can be used from any of several manufacturers. Specific items used here for which details are important include: |
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| CMA 12 microdialysis probes | CMA/ Microdialysis | CMA-12-XXX | These are available in various membrane lengths and cutoffs, indicated by specific codes in the ‘XXX.’ |
| Human insulin (Humulin) | Eli Lilly | N/a | |
| Liquid swivel | Instech | 375/D/22QM | This specific swivel has very low torque and internal volume, as well as a nonreactive quartz lining. |
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