Effect Of Propofol At Different Anesthesia Depth Onγ-aminobutyric Acid In Different Cerebral Regions In Dogs

y-aminobutyric acid (GABA), is the chief inhibitory neurotransmitter in mammalian central nervous system and acts synaptically to inhibit neurons.Many studies have showed that the release and transmit of neurotransmitters play a crucial role in the central nervous system(CNS), and the variations of propofol concentration in CNS have an effect on the release and transmit of neurotransmitters, which result in different anesthetic depth.Some studies concerning the relationship between propofol and GABA were carried out dating from the1980s. Propofol, which produces sedative and hypnotic effects in general anesthesia, may operate in inhibiting the synaptic transmission or down-regulating the excitation of the central nervous system. The possible mechanism of propofol is associated with the variable activity of ion channels via enhancing the effect of GABA and gated Cl-channel complexes which may play a critical role in general anesthesia. Orser revealed that the release of GABA in mouse neurons varied with different target-controlled infusion of propofol. The study showed low concentr ations(2-100umol/L) of propofol performed a concentration dependent enhancement of GABA whole cell current, extending the GABA-induced current decay time; mid-concentrations (100-2000umol/L) of propofol directly contributed to the GABA release of nerve endings, while high concentrations of propofol inhibited the release of GABA.Yao Shang-long found that clinically appropriate concentrations of propofol enhanced the effect of chloride ion uptake caused by GABA and GABAA receptor agonists (muscimol) in rat cortical brain slices, indicating that propofol promoted GABA and GABAA agonist mediated chloride channel activity. Besides it was likely to affect the GABAA receptor, via reducing the GABAA receptor channel inactivation and slowing down the desensitization process, resulting in the inhibitory postsynaptic potential (IPSP) recession delay.The research about cerebral uptake of propofol usually bases on the mass balance principles. By measuring the propofol concentrations in arterial and venous blood of cerebral circulation, we can calculate the cerebral concentrations and evaluate the cerebral uptake of propofol. Our previous study found that propofol distribution in different brain tissues in dogs was different:The propofol concentration was highest in the pons in light anesthesia group, and was higherest in the hypothalamus, lowest in the hippocampus in deep anesthesia group. The other studies showed that plasma propofol concentrations in the internal carotid artery and internal jugular vein reached equilibrium and propofol was distributed evenly except the dorsal thalamus contaning a higher concentration at30min of propofol constant infusion70mg.k g-1-h-1, propofol was distributed almost evenly among regional cerebral tissues at50min in dogs. The effect of propofol at different anesthesia depth on inhibitory or excitory neurotransmitter in different cerebral regions is still unknown.The chief aim of this study is to investigate the effect of propofol on GABA in different regions (dorsal thalamus, epithalamus, metathalamus, hypothalamus, subthalamus, frontal lobe, parietal lobe, temporal lobe, occipital lobe, hippocampus, cingulate gyrus, cerebellum, midbrain, pons, medulla oblongata, and cervical cord) when cerebral uptake and distribution of propofol reach equilibrium at different anesthesia depths.Material and methods1Animal preparation and groupingTwelve12-18-month-old healthy hybrid dogs weighing10-12kg were randomly divided into light anesthetic group(L group,n=6) and deep anesthetic group(D group, n=6).The experiment was scheduled during the day and raised in diet for10-hour were prior to experiment. The venous channel was established in the great saphenous vein of the right posterior limb.2Animal anesthesia and managementAll animals were anesthetized in light and deep anesthesia group with a single bolus dose of propofol (5.5and7.0mg.kg-1, respectively) in15s and followed by intervenous propofol infusion at a constant rate of55and70mg.kg-1.h-1respectively for50mins.After reached the appropriate depth of anesthesia, all animals were inserted endotracheal tube to keep PETCO230-38mmHg by mechanical ventilation (respiratory rate20-25breaths.min-1, tidal volume15ml.kg-1) and were monitored mean arterial pressure (MAP) and pulse rate (PR).3Sample collectionBlood samples were taken from the internal carotid artery and jugular vein respectively to measure plasma propofol concentrations50min after the start of the infusion in both groups. Then the dogs were sacrificed and cerebral tissues were taken from different regions (dorsal thalamus, epithalamus, metathalamus, hypothalamus, subthalamus, frontal lobe, parietal lobe, temporal lobe, occipital lobe, hippocampus, cingulate gyrus, cerebellum, midbrain, pons, medulla oblongata,and cervical cord) to measure GABA concentrations by high pressure liquid chromatography (HPLC). 4Sample processingAfter anticoagulant, each blood sample was centrifuged for10min at4℃centrifuge(3500r.min-1),200ul plasma was taken and placed in EP tube, after adding acetonitrile400ul,the sample was shocked in vortex device for2min,1000r.min-1centrifuge for10min.The supernatant was taken to analyzed.The brain tissues were extracted with acetonitrile(2ml.g-1) and homogenized for15min,the homogenates were placed in EP tube for centrifugaled for10min,10000r.min-1.The supernatant(50μl and200μl) for derivatization2min to analyzed immediately.5HPLC conditionsDetermination of propofol plasma concentrations:Column (Shim-pack VP-ODS,250nmx4.6mmID), Guard column(Shim-pack GVP-ODS,10mmx4.6mmID),mobile phase:15%pure water and85%methanol,the injection volume20ul,column temperature40℃,flow rate1ml.min-1,detection wavelength at270nm.Determining GABA in brain tissue:Column:Agilent XDB-C18(5μm,150mm×4.6mm),column temperature:35℃, mobile phase A:0.1mol.L-’sodium acetate, mobile phase B:methanol, excitation wavelength of330nm,emission wavelength, injection volume lOul.6Statistical analysisAll data were expressed as mean±standard deviation and analyzed with the Statistics Package for Social Sciences(SPSS,version13.0for windows;SPSS Inc,Chicago,IL,USA).Differences were considered statistically significant when P was less than0.05.We used one-way ANOVA,Paired-Samples T Test and Repeated Measure to test for differences. Multiple comparisons were analyzed by LSD test.Results1All animals reached objective anesthesia depths safely and quickly and there were no vomiting,asphyxia, apnea and other adverse reactions.The MAP and PR were significantly higher in group L than in group D(105.33±4.18vs89.50±1.87mmHg,90.50±3.62vs72.83±3.31bpm, respectively,P0.05).The PETCO2was36±1.41mmHg in group L and37±1.41mmHg in group D and there were no statistical differences(P>0.05).2The plasma propofol concentrations in internal carotid artery and jugular vein were no statistical differences in both group L (3.00±0.31and3.10±0.51ug.ml-1, resectively,t=0.335,P=0.751)and group D (6.41±0.05and6.40±0.11um.ml-1respectively,t=0.371,P=0.726).3GABA concentrations in cerebral regions were significantly higher in group D than in group L(P<0.05).The dorsal thalamus and hypothalamus showed greater GABA variations (83.83±2.23%and85.83±1.72%) compared to other cerebral regions at different anesthesia depths(P<0.05).Conclusions1In both groups, plasma propofol concentrations in the internal carotid artery and internal jugular vein reach equilibrium at50min of constant rate propofol infusion.2The variation of GABA is associated with the anesthesia depth of propofol. The GABA of dorsal thalamus, hypothalamus and their variations play an important role in propofol anesthesia.