Instrumentatie van Near-termijn Foetale Schapen voor Multivariate chronische niet-verdoofde Recordings
Summary
Chronisch geïnstrumenteerd niet-verdoofde foetale schapen model wordt toegepast om menselijke foetale ontwikkeling in gezondheid en ziekte te bestuderen, omdat het toestaat chirurgische plaatsing en onderhoud van catheters en elektroden herhaalde bloedafname, stof injectie, opname van bio-elektrische activiteit en in vivo imaging. We beschrijven de vereiste om dit model vast te stellen procedures.
Abstract
The chronically instrumented pregnant sheep has been used as a model of human fetal development and responses to pathophysiologic stimuli such as endotoxins, bacteria, umbilical cord occlusions, hypoxia and various pharmacological treatments. The life-saving clinical practices of glucocorticoid treatment in fetuses at risk of premature birth and the therapeutic hypothermia have been developed in this model. This is due to the unique amenability of the non-anesthetized fetal sheep to the surgical placement and maintenance of catheters and electrodes, allowing repetitive blood sampling, substance injection, recording of bioelectrical activity, application of electric stimulation and in vivo organ imaging. Here we describe the surgical instrumentation procedure required to achieve a stable chronically instrumented non-anesthetized fetal sheep model including characterization of the post-operative recovery from blood gas, metabolic and inflammation standpoints.
Introduction
Een verscheidenheid van dierlijke modellen bestaan voor de studie van normale en gecompromitteerd zwangerschappen, waaronder knaagdieren, primaten en huishoudelijke herkauwers. 1,2,3,4,5 Chronisch geïnstrumenteerde zwangere schapen is uitgebreid gebruikt voor 50 jaar een model van foetale ontwikkeling en reacties mens op pathofysiologische stimuli zoals lipopolysaccharide (LPS). 10/06 De laesies na blootstelling aan LPS nabootsen wat wordt gezien bij prematuren met periventriculaire leukomalacia, die als gevolg van vergelijkbaar rijpingsprocessen profiel van beide soorten. 11, 12
Andere zwangerschapscomplicaties hebben ook onderzocht in detail, zoals de ontdekking dat prenatale glucocorticoïden bevorderen longontwikkeling 13-15 en inzicht in de impact van Dysmaturiteit (IUGR) op de foetus 16,17.
Het uitgebreide gebruik van foetaal schapen model vanwege de uniqUE ontvankelijkheid van de niet-verdoofde foetaal schapen om de chirurgische plaatsing en onderhoud van catheters en elektroden, waardoor herhaalde bloedafname opnemen van bio-elektrische activiteit, de toepassing van elektrische stimulatie en in vivo beeldvorming van de hersenen. 18 Telemetrie is ook mogelijk, hoewel minder gebruikte maar vanwege de hogere verfijning te zetten als de aanvankelijke en onderhoudskosten. 19
Bovendien is de foetale schapen model is zeer veelzijdig vele variaties van instrumentatie zijn mogelijk afhankelijk van de maten van belang. Zo is het mogelijk voor opnemen dagen tot weken multivariate signalen in real time als foetale ademhalingsbewegingen elektrische hersenactiviteit, cardiovasculaire reacties, elektrocardiogram, doorbloeding een reeks organen gebruik stroming probes of microbolletjes, etc. Dankzij Deze veelzijdigheid, hebben een breed scala van studies uitgevoerd met inbegrip van de ontwikkeling van de Cardiovascular systeem 20,21, hypothalamus-hypofyse-bijnier (HPA) as 22, ontwikkeling van de hersenen ontwikkeling van 23 en slaap staten in het bijzonder 24, de effecten van hypoxie / asfyxie 25, therapeutische hypothermie 26, ontsteking 6-11, een combinatie van beide 27, glucocorticoïden 28,29, anti-depressiva 30, bronchopulmonale dysplasie (BPD) 31,32, foetale programmering 33,34,35,36,37,38,39 of de ontwikkeling van nieuwe foetale bewaking modaliteiten voor en tijdens de bevalling om maar een paar gebieden van onderzoek. 40,41,42,43
Het algemene doel van de gepresenteerde methode is om basisimplementatie dit veelzijdige model tonen. Het maakt vaststelling diverse acute en chronische experimentele protocollen bestuderen foetale fysiologie en pathofysiologie op de integratieve, orgaan, cellulair en moleculair niveau.
Protocol
Representative Results
Discussion
De anesthetische en chirurgische procedures worden die nodig zijn voor het opzetten van een diermodel voor de studie foetale fysiologie en pathofysiologie: chronisch geïnstrumenteerd niet-verdoofde foetaal schapen.
Vier kritische stappen in het protocol moet worden benadrukt. Ten eerste, langs de catheters en elektroden door het moederlijke flank: het is belangrijk dat dit tegelijkertijd gebeurt aan een inwendig orgaan letsel te voorkomen. Ten tweede, het veiligstellen van de uterotomy oper…
Offenlegungen
The authors have nothing to disclose.
Acknowledgements
authors gratefully acknowledge funding support from the Molly Towell Perinatal Research Foundation, Canadian Institutes of Health Research (CIHR), and Fonds de Recherche du Québec – Santé (FRQS) (to MGF) and CIHR-Quebec Training Network in Perinatal Research (QTNPR) (to LDD).
The authors wish to thank Esther Simard, Marco Bosa, Carl Bernard and Carmen Movila for technical assistance.
Materials
| ACE Light source | Schott-Fostec | A20500 | |
| Dissecting scissors | Fine Science Tools | 14060 – 11 | |
| Angled dissecting scissors | Fine Science Tools | 15006 – 09 | |
| Scalpel handle | Fine Science Tools | 10003 – 12 | alternating dissecting tool |
| Curved scalpel blades #12 | Fine Science Tools | 10012 – 00 | alternating dissecting tool |
| Bone scissors | Fine Science Tools | 16044 – 10 | |
| S & T suture tying forceps | Fine Science Tools | 00272 – 13 | |
| Dumont SS forceps – angled | Fine Science Tools | 11203 – 25 | |
| Braided silk suture size 6-0 | Teleflex Medical | 07 – 30 – 10 | |
| Medical Tape | transpore | 3M | |
| Ketamine hydrochloride 100 mg/ml | Hospira | NDC 0409 – 2051 – 05 | Final Does is 80 mg/kg |
| Tranqui Ved Injection (xylazine 100 mg/ml) | Vecdo | NDC 50989 – 234 – 11 | Final Does is 10 mg/kg |
| Reactive orange 14 | Sigma – Aldrich | R – 8254 | |
| Ringers Solution Components | Solution is gas equilibrated with 95% O2 and 5% Co2, final pH 7.4 | ||
| Sodium chloride | Sigma – Aldrich | S7653 | Final Concentration: 118 mM |
| Potassium chloride | Fisher Scientific | P217 – 3 | Final Concentration: 4.7 mM |
| Calcium chloride dihydrate | Fisher Scientific | C79 – 500 | Final Concentration: 2.5 mM |
| Potassium phosphate monobasic | Fisher Scientific | P -285 | Final Concentration: 1.2 mM |
| Magnesium sulfate | J.T. Baker | Jan-00 | Final Concentration: 0.57 mM |
| 4-(2-Hydroxyethyl)piperazine-1-ethanesulfonic acid (HEPES) | Fisher Scientific | BP 310 – 500 | Final Concentration: 5.95 g/L |
| Glucose | Sigma – Aldrich | G8270 | Final Concentration: 5.5 mM |
| LifeWindow | Digicare Biomedical Technology | ||
| CED bioamplifier and ADC units | Cambridge Electronic Design Limited, Unit 4, Science Park, Milton Road, Cambridge CB4 0FE ENGLAND. |
Bioamp: 1902; ADC: micro1401; Data acquisition software: Spike 2, V7.13 | |
| Neurolog analog signal bioamplifier | Digitimer Ltd 37 Hydeway Welwyn Garden City Hertfordshire, AL7 3BE, England |
NL108A | |
| ABL800Flex | Radiometer Canada; 200 Aberdeen Dr, London, ON N5V 4N2 | ||
| Eppendorf 5804R | Eppendorf Canada; 2810 Argentia Road, #2 Mississauga, Ontario, L5N 8L2 |
||
| Arrow Jugular Catheterization Set | Arrow International, Inc., 2400 Bernville Road, Reading, PA 19605 USA | ||
| Atravet | 10 mg/mL | ||
| Diazepam | 5mg/mL | ||
| Ketamine | Ketalar | 100 mg/mL | |
| Propofol | 10 mg/mL | ||
| SurgiVeT | Endotracheal Tubes; Smiths Medical ASD, Inc. St. Paul, MN 55112, USA | ||
| Cook Airway Exchange Catheter with RAPI-FIT Adapters | Cook Critical Care 750, Bloomington IN 47402-0489 USA | ||
| Dispomed Ventilator | Dispomed Ltd., 745 Nazaire-Laurin, Joliette, Quebec J6E 0L6 | ||
| BD Insyte-W | Becton Dickinson, Infusion Therapy Systems Inc., 9450 S State St, Sandy Utah 84070 USA | 22 to 20 G; 1 in [0.9 x 25 mm] to 1.16 in [1.1 x 30 mm] | |
| Edwards Lifesciences Ref: PX272 Pressure monitoring kit with TruWave Disposable Pressure | |||
| LifeWindow LW6000 | Digicare Biomedical Technology 107 Commerce Road, Boynton Beach, FL 33426-9365 USA | ||
| Gaymar | |||
| Babcock | |||
| Polyvinyl catheters | SCI (Scientific Commodities Inc.) | 2 meters | |
| 2-0 Vicryl | |||
| Castroviejo scissors | |||
| electrocardiogram (ECG) | LIFYY, Metrofunk Kabel-Union, Berlin, Germany | four copper electrodes in single sheath, 2 meters | |
| 2-O Vicryl | |||
| 3-0 Vicryl | |||
| PDS II USP | |||
| Trimethoprim sulfadoxine | |||
| Ampicillin | |||
| Stopcock | Argon Medical, Cat 041220001A | Double 4-way Stopcock with male luer lock | |
| Needles | Tyco Healthcare 8881202389 | Monoject aluminum hub blunt needles, 22Gx, 0.7mmx 38.1mm: for fetal arterial and venous catheters | |
| Needles | Tyco Healthcare 8881202322 | Monoject aluminum hub blunt needles, 16Gx, 1.6mmx38.1mm: for amniotic catheters |
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