in the annual report of the american association of poison control centers , around 5000 cases of intoxication with calcium channel blockers ( ccb ) were reported in 2013 , although this figure is over 11 000 when multi - drug ingestion is considered . more than half of these patients required hospitalization . there are studies reporting up to 10% ccb overdose mortality in children . verapamil and diltiazem are the most frequently used medications in ccb group . preparations of slow - prolonged release verapamil can be deadly , even in small quantities , especially in children . ccb overdose can lead to cardiovascular symptoms , such as hypotension , bradycardia , myocardial depression , as well as neurological ( e.g. , syncope and confusion ) , endocrinological ( e.g. , acidosis and hyperglycemia ) , and respiratory ( e.g. , respiratory depression and noncardiogenic pulmonary edema ) problems . standard treatment consists of gastrointestinal decontamination , high - dose insulin , calcium , vasopressors , and glucagon , although use of pacemakers , intra - aortic balloon pumps , extracorporeal membrane oxygenation , and venovenous hemodiafiltration has also been reported [ 59 ] . there are also animal studies and case reports that have reported successful use of lipid emulsion , beta - cyclodextrin , sugammadex ( a modified gamma - cyclodextrin ) , levosimendan , aminopyridine , and methylene blue [ 1017 ] . overdose of calcium channel blockers has high mortality and their use for suicide has been increasing . in this study our primary aim was to evaluate the effect of sugammadex , lipid emulsion , and sugammadex plus lipid emulsion on the survival time in a rat model of verapamil toxicity . our secondary aim was to investigate their effect on hemodynamic parameters in the same model . in this randomized , blinded , controlled trial , we compared the effect of sugammadex , lipid emulsion , and sugammadex plus lipid emulsion on hemodynamic parameters and survival time in a rat model of verapamil toxicity . the study was approved by yeditepe university school of medicine animal studies ethics board and was performed at yeditepe university school of medicine experimental research center in istanbul , turkey . twenty - eight sprague - dawley rats weighing 180250 grams were kept in a temperature ( 2224c ) and humidity - controlled environment with free access to food and water . during the experimental period , verapamil ( verapamil ratiopharm ; 5 mg/2 ml , ratiopharm gmbh , germany ) , sugammadex ( bridion ; 200 mg/2 ml , schering- plough , turkey ) , and 20% lipid emulsion ( clinoleic 500 ml , eczacba - baxter , turkey ) were purchased for use in this study . all rats weights and ages were noted and rats were randomly separated into 4 groups : control group ( group c ) , sugammadex group ( group s ) , lipid emulsion group ( group l ) , and lipid emulsion plus sugammadex group ( group sl ) . rats were anesthetized using 60 mg / kg intraperitoneal ketamine . following a period of 611 minutes for hemodynamic stabilization and depth of anesthesia , rats were placed on a warm surface ( 38c ) and 1 l / m o2 was administered . after electrocardiographic ( ecg ) monitoring via alligator clips , the right femoral area was surgically dissected and p50 cannulas were placed in the femoral artery and vein . to counteract any blood loss up to this time , 1 ml of saline solution was administered . for medication and fluid infusions , a cma 400 syringe pump ( cma microdialysis , kista , sweden ) ecg and mean arterial pressure ( map ) measurements were performed using lab chart power lab 8/30 ( ad instruments , new south wales , australia ) . we administered 37.5 mg / kg / hr dose of verapamil ( 2.5 mg / ml ) via the femoral vein to all rats . t0 was the time when the infusion started , and infusion continued until death was confirmed . death was accepted as asystole on ecg or no respiratory effort for over 1 minute . in group c , 5 minutes after beginning of verapamil infusion , rats were given 12.4 ml / kg normal saline for a period of 5 minutes . this amount represents a 20% volume based on an average rat blood volume of 62 ml / kg . in group s , following a 16 mg / kg sugammadex bolus , saline was given to complete the total dose of 12.4 ml / kg . in group l , 20% intralipid emulsion was administered at 12.4 ml / kg . in group sl , following 16 mg / kg sugammadex bolus , 20% lipid emulsion was administered in an amount to complete a total of 12.4 ml / kg . the data recorder who noted infusion times , time of death , total verapamil dosage ( mg / kg ) , mean arterial pressures , and heart rate information was blinded to group identity . statistical analysis was performed using ibm spss statistics 22 ( ibm spss , turkey ) . the kolmogorov - smirnov test was used to evaluate the distribution of data and parameters were found not to show normal distribution . parameters were compared using the kruskal - wallis test and mann - whitney u test . in this randomized , blinded , controlled trial , we compared the effect of sugammadex , lipid emulsion , and sugammadex plus lipid emulsion on hemodynamic parameters and survival time in a rat model of verapamil toxicity . the study was approved by yeditepe university school of medicine animal studies ethics board and was performed at yeditepe university school of medicine experimental research center in istanbul , turkey . twenty - eight sprague - dawley rats weighing 180250 grams were kept in a temperature ( 2224c ) and humidity - controlled environment with free access to food and water . during the experimental period , verapamil ( verapamil ratiopharm ; 5 mg/2 ml , ratiopharm gmbh , germany ) , sugammadex ( bridion ; 200 mg/2 ml , schering- plough , turkey ) , and 20% lipid emulsion ( clinoleic 500 ml , eczacba - baxter , turkey ) were purchased for use in this study . all rats weights and ages were noted and rats were randomly separated into 4 groups : control group ( group c ) , sugammadex group ( group s ) , lipid emulsion group ( group l ) , and lipid emulsion plus sugammadex group ( group sl ) . rats were anesthetized using 60 mg / kg intraperitoneal ketamine . following a period of 611 minutes for hemodynamic stabilization and depth of anesthesia , rats were placed on a warm surface ( 38c ) and 1 l / m o2 was administered . after electrocardiographic ( ecg ) monitoring via alligator clips , the right femoral area was surgically dissected and p50 cannulas were placed in the femoral artery and vein . to counteract any blood loss up to this time , 1 ml of saline solution was administered . for medication and fluid infusions , a cma 400 syringe pump ( cma microdialysis , kista , sweden ) ecg and mean arterial pressure ( map ) measurements were performed using lab chart power lab 8/30 ( ad instruments , new south wales , australia ) . we administered 37.5 mg / kg / hr dose of verapamil ( 2.5 mg / ml ) via the femoral vein to all rats . t0 was the time when the infusion started , and infusion continued until death was confirmed . death was accepted as asystole on ecg or no respiratory effort for over 1 minute . in group c , 5 minutes after beginning of verapamil infusion , rats were given 12.4 ml / kg normal saline for a period of 5 minutes . this amount represents a 20% volume based on an average rat blood volume of 62 ml / kg . in group s , following a 16 mg / kg sugammadex bolus , saline was given to complete the total dose of 12.4 ml / kg . in group l , 20% intralipid emulsion was administered at 12.4 ml / kg . in group sl , following 16 mg / kg sugammadex bolus , 20% lipid emulsion was administered in an amount to complete a total of 12.4 ml / kg . the data recorder who noted infusion times , time of death , total verapamil dosage ( mg / kg ) , mean arterial pressures , and heart rate information was blinded to group identity . statistical analysis was performed using ibm spss statistics 22 ( ibm spss , turkey ) . the kolmogorov - smirnov test was used to evaluate the distribution of data and parameters were found not to show normal distribution . parameters were compared using the kruskal - wallis test and mann - whitney u test . age , weight , temperature , and basal respiratory rates were similar for all groups ( p>0.05 ) . average time to death was 21.09.57 minutes for group c , 35.5710.61 minutes for group s , 37.1416.6 minutes for group l , and 49.8627.56 minutes for group sl . when inter - group comparison was performed , the time to death was significantly shorter in group c ( p<0.05 ) . l or sl . similarly , while the average lethal dose was significantly different in group c when compared to other groups ( p<0.05 ) , there was no difference between groups s , l , or sl ( p>0.05 ) ( table 1 ) . figure 1 shows the box plot graphic for time to death , figure 2 shows the kaplan - meier analysis , and figure 3 shows the relation between administered dose and number of living rats . when time to death data was analyzed , rats in group sl were found to live longer than other groups . while only 1 rat survived longer than 30 minutes in group c , only 2 rats died before 30 minutes in the remaining groups . the number of rats died before 20 minutes was 4 for group c , 1 each for group l and sl , and none in group s ( figures 1 , 2 ) . tables 2 and 3 show average heart rate and map at the time when verapamil infusion was commenced ( t0 ) , the time when drugs and/or saline infusion were started ( t5 ) , and the time when infusion was discontinued ( t10 ) . while there was no difference in heart rates at t0 or t5 between groups , at t10 heart rates of groups s and sl were significantly higher compared to groups c and l ( p<0.05 ) ( table 2 ) . no difference was observed between groups at t0 , t5 , or t10 for map ( p>0.5 ) ( table 3 ) . heart rates and average map from t0 until asystole is shown in figures 4 and 5 . heart rates and average map from t0 however , a slight increase in heart rate and map was seen after sugammadex infusion in group s , and it was stable for longer than in other groups ( figures 3 , 4 ) . when compared with the second group , in which sugammadex was administered , heart rate and map were slightly higher in group sl than in groups c and l. ecg analysis showed sinus bradycardia to be the dominant rhythm . following asystole , most rats were observed to have some respiratory effort in the form of gasping . treatment generally involves gastric decontamination and symptomatic treatment to counteract the drug s negative effects [ 35 ] . research continues on drugs that can be used in verapamil toxicity . in intoxications caused by lipophilic drugs , a study performed in 2003 found that 20% intralipid administered to dogs after bupivacaine - induced cardiac arrest increased the success rate of resuscitation . in 2006 , tebbutt et al .   found that intravenous infusion of 20% lipid emulsion increased time to death and ld50 in a rat model of verapamil toxicity . more recently , several case reports and series reported beneficial effects of intravenous lipid emulsion in serious verapamil overdose [ 5,2024 ] . lipid sink  theory is the most accepted explanation for why intravenous lipid emulsion seems to be effective in verapamil and other lipophilic drugs [ 13,2224 ] . there are some studies on the effect of cyclodextrins in verapamil toxicity [ 1618 ] . mottram et al .   demonstrated prolonged survival in a verapamil toxicity model when sulfobutylether--cyclodextrin ( sbe - cd ) , an agent known to bind to verapamil , was administered . ozbilgin et al .   reported that sugammadex , a modified gamma cyclodextrin , at 16 mg / kg dose , prolonged survival and increased ld in rats when administered for verapamil toxicity , similar to intralipid and sbe - cd . it is known to encapsulate steroidal non - depolarizing muscle relaxants used for anesthesia in order to reverse the effect of these drugs . although it encapsulates verapamil 120700 times less than rocuronium , its beneficial effects have been shown for verapamil toxicity . our study found that when compared to a control group , the administration of intravenous lipid , sugammadex , or both prolonged survival and increased verapamil ld in animals . however , there was no statistically significant difference between survival or ld among groups s , l , and sl . when other groups were compared with group sl for mean survival time , there was a 2.5-fold increase for group c , and a nearly 1.5-fold increase for groups s and l. there is a correlation between our results and those reported by ozbilgin et al .   in terms of survival and verapamil ld in the sugammadex ( 16 mg / kg ) group . compared to our results , tebbutt et al .   reported survival times that were 4 minutes longer for the control group and 8 minutes longer in the lipid group . this difference could be due to the difference in type of animal and dosage of intraperitoneal ketamine . in our study the difference in the brand and content of these 2 lipid solutions could also explain the difference in survival . compared these 2 types of lipid emulsions in local anesthetic toxicity and found no significant difference . apart from comparing survival between groups , we also monitored the changes in heart rate and arterial blood pressure during verapamil infusion to determine the requirement for positive inotropic agents and , if necessary , their timing . when compared to other groups , there was a pronounced difference for group s and a less pronounced difference for group sl for heart rate and map . when this data is taken into account , the administration of sugammadex in verapamil overdose could lead to lower requirement for inotropic agents . similarly , ozbilgin et al .   found that heart rate decrease was least in the sugammadex 16 mg / kg group . because sugammadex does not seem to affect hemodynamics when used to reverse the effect of steroidal muscle relaxants in anesthesia , this effect could be due to the rapid binding of sugammadex to verapamil in the circulatory system . however , our data suggest that less positive inotropic agents are be required when only sugammadex or sugammadex plus 20% lipid emulsion is infused for verapamil overdose . the suggested dosage for sugammadex , which this study and a previous study   has demonstrated to increase survival for verapamil toxicity in rats , is 14 mg / kg doses for reversing the effect of nondepolarizing muscle relaxants , although doses of 81632 mg / kg can be used for reversal deep neuromuscular blockage . in accordance with data from ozbilgin et al . that shows efficacy for verapamil toxicity in rat model , we used 16 mg / kg dosage in this study . although our data has demonstrated that these doses increase survival in rats and keep hemodynamic parameters more stable for a longer time , most possibly through its effect on binding verapamil . however , further studies are required to determine the optimum dosage for its clinical use in humans . the reported dosage of 20% lipid emulsion for rat models of verapamil toxicity is 12.4 ml / kg . however , its usage in human toxicity is different and it is generally used as a 24-hour or longer continuous infusion following a bolus dose . there are no verapamil toxicity rat models with lipid interventions as a continuous infusion following a bolus . several studies have demonstrated the effect of lipid emulsion for verapamil toxicity in humans . in our study we compared the use of sugammadex alone , lipid emulsion alone , and the combination of sugammadex and lipid emulsion . although we found that survival was increased in all groups when compared to the control group , there was no statistically significant difference for the combination of sugammadex and lipid emulsion . it is possible that intraperitoneal ketamine used for anesthesia in rats could lead to hemodynamic changes . however , procedures such as insertion of femoral artery and venous cannula , plus the requirement for rats to stay immobile during infusion , lead to the necessity of anesthesia . although this study was planned to include mechanical ventilation support with blood gasses and blood glucose monitoring throughout all procedures , technical difficulties and financial limitations did not make this possible . however , if we do not take into account any effect of bradypnea , and considering that the last event for all rats was respiratory arrest , we do not believe that mechanical ventilation would have changed the end results . we were not able to determine the effect of early complications leading to death in this study . we could not exclude rats that had adverse effect to intravenous lipid emulsion , such as hypersensitivity reaction or pulmonary or cerebral fat emboli in groups l and sl [ 10,2123,28 ] . sprague - dawley rats have no or at most a very short calcium - dependent phase ii action potential plateau and may not reproduce the cardiotoxic effects of ccbs seen in humans . treatments found to have good efficacy in other animal models ( such as high - dose insulin ) were not tested . monitoring could have been improved by including pressure - volume assessment to estimate lv contractility , and blood verapamil concentrations . although there are reports on the use of lipid emulsion in verapamil toxicity in humans , there are no reports for the use of sugammadex . therefore , further studies are required to demonstrate the effectiveness and optimal dosage of sugammadex for verapamil toxicity in humans , as well as evaluating the combination of sugammadex and lipid emulsion . it is possible that intraperitoneal ketamine used for anesthesia in rats could lead to hemodynamic changes . however , procedures such as insertion of femoral artery and venous cannula , plus the requirement for rats to stay immobile during infusion , lead to the necessity of anesthesia . although this study was planned to include mechanical ventilation support with blood gasses and blood glucose monitoring throughout all procedures , technical difficulties and financial limitations did not make this possible . however , if we do not take into account any effect of bradypnea , and considering that the last event for all rats was respiratory arrest , we do not believe that mechanical ventilation would have changed the end results . we were not able to determine the effect of early complications leading to death in this study . we could not exclude rats that had adverse effect to intravenous lipid emulsion , such as hypersensitivity reaction or pulmonary or cerebral fat emboli in groups l and sl [ 10,2123,28 ] . sprague - dawley rats have no or at most a very short calcium - dependent phase ii action potential plateau and may not reproduce the cardiotoxic effects of ccbs seen in humans . treatments found to have good efficacy in other animal models ( such as high - dose insulin ) were not tested . monitoring could have been improved by including pressure - volume assessment to estimate lv contractility , and blood verapamil concentrations . although there are reports on the use of lipid emulsion in verapamil toxicity in humans , there are no reports for the use of sugammadex . therefore , further studies are required to demonstrate the effectiveness and optimal dosage of sugammadex for verapamil toxicity in humans , as well as evaluating the combination of sugammadex and lipid emulsion . this study demonstrated that rat survival in verapamil toxicity is prolonged with the application of intravenous lipid emulsion and sugammadex separately or as combined treatment . in all cases , the cardiotoxic effect of verapamil the administration of sugammadex at 16 mg / kg alone or combined with 20% intravenous lipid emulsion could lead to a lessened requirement for positive inotropic agents .
backgroundtoxicity of calcium channel blockers leads to high patient mortality and there is no effective antidote . 
 the benefit of using 20% lipid emulsion and sugammadex has been reported . 
 the present study measured the effect of sugammadex and 20% lipid emulsion on hemodynamics and survival in a rat model of verapamil toxicity.material/methodsin this single - blinded randomized control study , rats were separated into 4 groups of 7 rats each : sugammadex ( s ) , sugammadex plus 20% lipid emulsion ( sl ) , 20% lipid emulsion ( l ) , and control ( c ) . 
 heart rates and mean arterial pressures were monitored and noted each minute until death.resultsaverage time to death was 21.09.57 minutes for group c , 35.5710.61 minutes for group s , 37.1416.6 minutes for group l and 49.8627.56 minutes for group sl . 
 time to death was significantly longer in other groups than in the control group ( p<0.05).conclusionsverapamil overdose is has a comparatively high mortality rate and there is no effective antidote . 
 treatment generally involves gastric decontamination and symptomatic treatment to counteract the drug s negative effects . in animal studies sugammadex and lipid emulsion 
 had a positive effect on survival in patients with calcium channel blocker toxicity . 
 sugammadex and intralipid increased survival in a rat model of verapamil toxicity . 
 the combination of both drugs may decrease cardiotoxicity . 
 sugammadex alone or combined with 20% lipid emulsion reduce the need for inotropic agents . 
 the mechanism requires clarification with larger studies .