Platelets play a variety of important roles in the body, most notably in hemostasis. They are essential to primary hemostasis, or the formation of a hemostatic plug, and are also important for secondary hemostasis. Primary hemostasis and secondary hemostasis have 3 overlapping phases called initiation, amplification, and propagation. Once clot formation is initiated via tissue factors, subsequent thrombin formation activates several clotting factors, including V, VIII, and XI, and stimulates platelets to undergo shape and surface changes, activate surface receptors, and release granular contents that recruit and promote activation of additional platelets. Platelets also provide a phospholipid surface for the accumulation of procoagulant complexes, thereby propagating and further amplifying the hemostatic process.1 Although thrombin is recognized as the most potent platelet agonist, ADP and TXA2 also mediate platelet activation via specific transmembrane receptors on the platelet surface.
Acepromazine maleate is a phenothiazine neuroleptic agent that can inhibit platelet function.2–5,a One proposed mechanism of inhibition involves reduced platelet aggregation in response to ADP.2 Interference with AA or membrane lipid bilayer disorganization has also been implicated.3,4 These observations have resulted in recommendations to avoid the use of acepromazine in patients with coagulation disorders or in which severe hemorrhage is expected.2,a Acepromazine is used as a premedicant in a variety of research settings, including the evaluation of coagulation variables.6–10 Thus, understanding the effects of acepromazine on hemostatic function is important for accurate interpretation of results of diagnostic evaluations.
Thromboelastography is emerging in veterinary medicine as a useful tool in the overall assessment of thrombin-based clot formation.11–17 Thrombin is a major product of kaolin-activated thromboelastography, and because thrombin is such a potent platelet activator, the contribution of individual and potentially weaker platelet agonists may be obscured by thrombin activity. For other platelet agonists to be assessed, the formation of thrombin must be bypassed. This can be accomplished by the addition of reptilase, a purified thrombin-like enzyme found in snake venom, and activated factor XIII to a heparinized blood sample in lieu of standard activation.18 Reptilase cleaves fibrinogen into fibrin and, with activated factor XIII, creates a cross-linked fibrin mesh without the formation of thrombin.19 The contribution of specific agonists, such as ADP or AA, on platelet activation can then be individually assessed. Clopidogrel bisulfate exerts antiplatelet effects via inhibition of the P2Y12 receptor,20 whereas acetylsalicylic acid inhibits AA-induced platelet aggregation via inhibition of cyclooxygenase and cyclooxygenase's downstream production of thromboxane.21 By use of a 3-tiered modification of thromboelastography, the efficacy of clopidogrel and acetylsalicylic acid treatment can be assessed because only those platelets not inhibited will be able to respond to the addition of ADP and AA, respectively.20,21 This modification of thromboelastography for evaluating platelet function is performed by use of a thromboelastography analyzer.b
The purpose of the study reported here was to evaluate the effects of IV administration of acepromazine on the function of platelets when stimulated with ADP or AA in healthy dogs. We hypothesized that the administration of acepromazine to healthy dogs at therapeutic doses would not affect platelet function, as assessed by use of a modified thromboelastography assay.
Materials and Methods
Animals—Six adult mixed-breed dogs (mean body weight, 21 kg; range, 7 to 30 kg) maintained as part of a teaching and research colony at the university were used for the study. Five of the dogs were spayed females, and 1 was a sexually intact female that was not in estrus. All dogs were housed in the research facility at North Carolina State University throughout the study. Each dog was judged to be healthy on the basis of results of a physical examination, CBC, serum biochemical analysis, and a coagulation panel (consisting of assessments of prothrombin time, activated partial thromboplastin time, and d-dimer and fibrinogen concentrations) performed on citrated plasma. The study was approved by the North Carolina State University Institutional Animal Care and Use Committee.
Study design—The study had a 3-way crossover design that was conducted in a random order. Each dog was assigned a number, and treatment order (saline solution, LDA, and HDA) was determined by random selection out of a hat. Food, but not water, was withheld from dogs for approximately 12 hours prior to each treatment. There was a minimum 3-day washout period between treatments. The same investigator (BJC) performed all tests and was aware of the treatments administered.
Treatment protocols—Treatments were as follows: saline (0.9% NaCl) solution (control; 1 to 2 mL, IV, once), LDAc (0.05 mg/kg, IV, once), and HDA (0.1 mg/kg, IV, once). Saline solution or acepromazine was administered as a single bolus directly into a cephalic vein with a 3-mL syringe and 22-gauge needle. Confirmation of IV administration was made by aspirating a small volume of blood back into the syringe prior to injection and lack of evidence of perivascular swelling following injection.
Blood sample collection—Blood samples were collected immediately before (0 minutes) treatment and at 30 and 240 minutes after treatment administration. Each dog was positioned in lateral recumbency, and a blood sample was collected from a jugular vein directly into evacuated tubes by use of a 21-gauge butterfly catheter.d Two to 3 mL of blood was collected into an evacuated tubee containing no additives before filling a 1.8-mL tubef containing sodium citrate and a 4-mL tubeg containing lithium heparin. All samples were collected by the same investigator (BJC), and the jugular vein from which samples were obtained was alternated between venipunctures; for the third sample, the venipuncture site was cranial to the previous venipuncture site. Additionally, each phlebotomy was assigned a difficulty score from 0 to 3 as follows: 0 = venipuncture was successfully performed with 1 attempt, no readjustments of the needle within the vessel were made, and there was constant blood flow into the evacuated tube; 1 = venipuncture was successfully performed with 1 attempt with slight readjustment and nearly constant flow of blood into the evacuated tube; 2 = 1 or 2 attempts at venipuncture were performed, multiple readjustments in positioning of the needle were required, or there was moderate interruption in blood flow into the evacuated tube; and 3 = numerous attempts at venipuncture or considerable repositioning of the needle within the vessel were performed, or there was marked interruption of blood flow into the evacuated tube.
Modified thromboelastography—Four thromboelastography assays, consisting of TEGthrombin, TEGfibrin, TEGADP, and TEGAA, were performed on blood samples collected before (0 minutes [baseline]) and 30 and 240 minutes after treatment (saline solution, LDA, or HDA). Blood samples were kept at 23°C for 30 minutes after phlebotomy. Subsequently, all assays were performed simultaneously. Two analyzers (4 channels) were used, and the same assays were performed on the same channel at 37°C for every assay. Each test was performed on a commercially available thromboelastography analyzerh by use of a kit for modified thromboelastographyi provided by the manufacturer; the kit was stored in a refrigerator at 4°C until 30 minutes prior to testing, at which time it was removed and allowed to warm to room temperature (approx 23°C). Each kit contained the reagents needed for test completion (reptilase, ADP, AA, and kaolin). Reagents were prepared according to the manufacturer's instructionsi just prior to test performance for each assay. For TEGthrombm, 1 mL of blood from the tube containing sodium citrate was transferred into a tube containing a kaolin activatorj and inverted 5 times. Immediately thereafter, 0.34 mL of kaolin-activated blood was pipetted into the thromboelastography cup, which contained 20 μL of calcium chloride,k and the test was initiated. For TEGfibrin, 10 μL of reptilase reagenti (containing reptilase and factor XIII) was pipetted into the thromboelastography cup, followed by the addition of 0.36 mL of blood from the tube containing lithium heparin; the combination was mixed and the test was initiated. For TEGADP and TEGAA, 30 μL of the ADP reagent and 20 μL of the AA reagent were added to thromboelastography cups, respectively, in addition to 10 μL of the reptilase reagent. For each test, 0.36 mL of blood from the tube containing lithium heparin was added to the thromboelastography cup and mixed by gently swirling the sample 3 times with the pipette tip before the tests were initiated. The final concentrations of ADP and AA were 5.7μM and 1.9mM, respectively. All remaining doses and procedures were in accordance with the manufacturer's instructions.i Assays were terminated once data for R (ie, reaction time), K (ie, clot formation time), α angle, and MA were obtained.
Determination of ADP and AA concentrations—The amounts of ADP and AA reagents used in the modified thromboelastography assay in the study reported here were selected on the basis of preliminary data obtained from 6 healthy mixed-breed dogs. The dogs were deemed healthy on the basis of history and results of physical examinations, CBCs, and serum biochemical analyses. Modified thromboelastography was performed as described, with the exception that the activator (ADP or AA) was added in 10-, 20-, or 30-μL aliquots. These amounts were selected on the basis of the volume of activator provided by the manufacturer and the volume of solution that could be contained within the thromboelastography cup without overfilling the cup. The optimal amount of activator was determined to be that at which no further consistent increase in MA was achieved or the maximum volume (30 μL) that could be contained in addition to the blood sample within the thromboelastography cup.
Data and statistical analysis—Changes in overall coagulation were evaluated by comparing baseline (0 minutes) TEGthrombin values for R, K, α angle, and MA to those values obtained at 30 and 240 minutes after administration of each treatment (saline solution, LDA, and HDA). At each time, changes in platelet activation were evaluated by comparing the MA determined via TEGthrombin with the MA determined via TEGADP and the MA determined via TEGAA, respectively. For each treatment at a given time, the change in MA as a result of ADP activation or AA activation was calculated by dividing the MA determined via TEGADP and TEGAA, respectively, by the MA determined via TEGthrombin (ie, change in TEGADP MA was the TEGADP MA divided by TEGthrombin MA and change in TEGAA MA was the TEGAA MA divided by TEGthrombin MA). Change in platelet activation was further assessed by comparing the percentage inhibition in MA as described22 by use of the following formulas:
Because the data did not follow assumptions for a parametric distribution, nonparametric tests were used for comparisons. Additionally, although minimal, there were missing data points, so the Skillings-Mack test was used for each comparison.23 The Skillings-Mack test is an extension of the Friedman test for nonparametric repeated-measures analysis and is equivalent to the Friedman test when no data are missing. The Skillings-Mack test is also valid in the case of ties (equal ranks),23 which happened frequently in the present study. Analyses were stratified by time and treatment to test for differences across time and treatment.
With few exceptions, P values were determined by use of a χ2 distribution for each comparison. If the sample size is small, simulations are preferable for obtaining more accurate P values because the P value from the χ2 approximation is likely to be conservative.24 If there were ties (equal ranks), mean ranks were assigned. The Skillings-Mack statistic can be calculated when there are ties; however, the P value calculated from the assumed null χ2 distribution becomes more conservative as the number of ties increases. To provide a more accurate P value, simulations (by data shuffling permutations) were used to approximate the distribution of the Skillings-Mack statistics under the null hypothesis and were conditional on the particular missing-data structure and tied rankings. To preserve the missing-data structure, sorting on random numbers was not applied when there were missing data. This method was used to determine the raw P values for each comparison.
In an effort to improve the power of the study reported here, data on the difficulty of obtaining blood samples in the present study were pooled with similar data from another studyl conducted concurrently by our laboratory group that investigated the association of site of blood sample collection and technique with thromboelastography results in healthy dogs. Data on the difficulty of obtaining blood samples via the same venipuncture technique from that studyl were pooled with the data obtained in the present study, and differences were tested by use of a repeated-measures ANOVA.
To correct for the number of comparisons performed in the study, a Bonferroni correction was applied to control the overall family-wise error rate of the experiment; P values were multiplied by the total number of tests so that the family-wise type I error rate was 0.05. The corrected P values were evaluated, and corrected values of P < 0.05 were considered significant for all analyses.
To aid in the interpretation of the study results, power calculations by use of the Bonferroni-corrected type I error rate and parametric assumptions for repeated-measures ANOVA were performed to assess the power to detect changes in MA. All data were analyzed and power calculations were performed with commercially available software.m
Results
For the TEGthrombin assay, values for R, MA, K, or α angle did not differ significantly among the treatments (saline solution, LDA, and HDA) at any time (Table 1). At each respective time, there was no significant difference in the change in MA determined via TEGADP, change in MA determined via TEGAA (Figure 1), percentage inhibition for TEGADP, or percentage inhibition for TEGAA among treatments. Also, there was no significant association between the difficulty of obtaining a blood sample and the change in MA for any blood sample obtained or assay performed (determined on the basis of pooled data; 52 blood samples obtained from the present study and 32 blood samples obtained from the concurrent studyl).
Median and interquartile range for percentage change in TEGADP MA (A) and percentage change in TEGAA MA (B) in blood samples collected immediately before (0 minutes) and 30 and 240 minutes after administration of saline (0.9% NaCl) solution (1 to 2 mL, IV; white bars), LDA (0.05 mg/kg, IV; gray bars), or HDA (0.1 mg/kg, IV; black bars) to 6 healthy adult mixed-breed dogs. Each dog received each treatment with a minimum 3-day washout period between treatments. Change in TEGADP MA was calculated as TEGADP MA divided by TEGthrombin MA, and change in TEGAA MA was calculated as TEGAA MA divided by TEGthrombin MA.
Citation: American Journal of Veterinary Research 73, 5; 10.2460/ajvr.73.5.595
Median and interquartile range for percentage change in TEGADP MA (A) and percentage change in TEGAA MA (B) in blood samples collected immediately before (0 minutes) and 30 and 240 minutes after administration of saline (0.9% NaCl) solution (1 to 2 mL, IV; white bars), LDA (0.05 mg/kg, IV; gray bars), or HDA (0.1 mg/kg, IV; black bars) to 6 healthy adult mixed-breed dogs. Each dog received each treatment with a minimum 3-day washout period between treatments. Change in TEGADP MA was calculated as TEGADP MA divided by TEGthrombin MA, and change in TEGAA MA was calculated as TEGAA MA divided by TEGthrombin MA.
Citation: American Journal of Veterinary Research 73, 5; 10.2460/ajvr.73.5.595
Median and interquartile range for percentage change in TEGADP MA (A) and percentage change in TEGAA MA (B) in blood samples collected immediately before (0 minutes) and 30 and 240 minutes after administration of saline (0.9% NaCl) solution (1 to 2 mL, IV; white bars), LDA (0.05 mg/kg, IV; gray bars), or HDA (0.1 mg/kg, IV; black bars) to 6 healthy adult mixed-breed dogs. Each dog received each treatment with a minimum 3-day washout period between treatments. Change in TEGADP MA was calculated as TEGADP MA divided by TEGthrombin MA, and change in TEGAA MA was calculated as TEGAA MA divided by TEGthrombin MA.
Citation: American Journal of Veterinary Research 73, 5; 10.2460/ajvr.73.5.595
Median (range) for TEGthrombin assays and MA results determined via TEGADP and TEGAA in blood samples obtained immediately before (0 minutes) and 30 and 240 minutes after administration of saline (0.9% NaCl) solution (1 to 2 mL, IV), LDA (0.05 mg/kg, IV), or HDA (0.1 mg/kg, IV) to 6 healthy adult mixed-breed dogs; each dog received each treatment with a minimum 3-day washout period between treatments.
| Time (min) | ||||
|---|---|---|---|---|
| Variable | Treatment | 0 | 30 | 240 |
| TEGthrombin R (min) | Saline solution | 5.6 (4.7–7.1) | 6.2 (5.1–7.5) | 5.7 (3.2–6.8) |
| LDA | 5.1 (4.7–6.5) | 4.7 (3.3–5.9) | 5.8 (1.8–7.0) | |
| HDA | 4.5 (2.5–6.2) | 5.4 (3.1–6.1) | 6.4 (4.7–8.6) | |
| TEGthrombin K (min) | Saline solution | 2.1 (1.5–2.6) | 2.0 (1.5–2.7) | 2.0 (1.3–2.1) |
| LDA | 1.9 (1.5–2.2) | 1.4 (1.3–1.8) | 1.7 (1.1–2.2) | |
| HDA | 1.7 (0.9–1.9) | 1.7 (1.2–1.8) | 1.9 (1.7–2.6) | |
| TEGthrombin α angle (°) | Saline solution | 61.5 (55.1–67.8) | 62.4 (54.4–69.0) | 61.3 (59.7–71.0) |
| LDA | 64.5 (59.0–67.0) | 68.9 (63.9–70.2) | 64.0 (60.2–74.7) | |
| HDA | 66.5 (63.0–76.0) | 67.0 (46.1–71.8) | 63.2 (54.2–64.0) | |
| TEGthrombin MA (mm) | Saline solution | 55.1 (51.4–65.9) | 56.6 (52.0–69.2) | 61.3 (58.2–69.2) |
| LDA | 58.8 (58.0–64.2) | 60.1 (57.0–67.4) | 61.2 (57.6–64.1) | |
| HDA | 60.7 (55.9–67.6) | 63.0 (60.1–66.6) | 59.8 (57.6–66.4) | |
| TEGADP MA (mm) | Saline solution | 38.3 (27.2–51.3) | 43.8 (22.7–54.3) | 51.2 (20.5–51.8) |
| LDA | 32.5 (21.0–49.2) | 50.8 (27.5–61.4) | 53.0 (36.7–55.5) | |
| HDA | 30.9 (18.8–51.1) | 49.5 (26.9–57.8) | 52.0 (38.6–60.7) | |
| TEGAA MA (mm) | Saline solution | 23.1 (20.5–51.5) | 38.8 (16.3–53.6) | 45.7 (19.7–57.6) |
| LDA | 31.3 (16.5–46.3) | 41.5 (26.3–55.1) | 52.2 (23.3–53.9) | |
| HDA | 26.4 (15.8–47.3) | 44.0 (36.0–51.5) | 50.4 (39.4–60.1) | |
K = Clot formation time. R = Reaction time.
For the present study, calculations revealed that there was 80% power to detect differences ≥ 1.72 mm for the change in MA determined via TEGADP and differences ≥ 1.75 mm for the change in MA determined via TEGAA. For the purposes of the power calculations, a change of 5 mm in the MA for TEGADP or TEGAA was considered clinically relevant by the authors. The present study had a power of > 99% to detect MA changes as low as 5 mm by both TEGADP and TEGAA.
Discussion
In the study reported here, a single dose of acepromazine at 0.05 or 0.1 mg/kg administered IV did not affect in vitro thrombin-mediated hemostasis or platelet function as determined by platelet activation with ADP or AA via a modified thromboelastography assay in blood samples collected from healthy dogs. These results differ from those of studies2,5,a in dogs and humans that were conducted to evaluate the effects of phenothiazines on platelet aggregation. Following sedation with acepromazine and atropine sulfate in female adult Beagles, there was decreased platelet aggregation as determined by use of whole-blood aggregometry.2 However, when these dogs2 were anesthetized and underwent surgery, no clinically relevant bleeding was observed. The investigators concluded that although dogs with clinically normal clotting capabilities did not have altered hemostasis, acepromazine may exacerbate hemorrhage in dogs with underlying platelet abnormalities.2 In another study,a conducted to evaluate the effects of acepromazine on platelet counts and aggregation (the type of aggregometer used was not specified) in dogs, the administration of acepromazine (0.1 mg/kg, IM) resulted in significant decreases in platelet counts and the percentage of platelet aggregation, but no significant changes in clotting and bleeding times were observed. A study5 conducted to evaluate the effects of various drugs on platelet aggregation and bleeding times in humans revealed that chlorpromazine hydrochloride did not cause a change in bleeding times or alter ADP-induced turbidimetric platelet aggregation, but it did cause decreased epinephrine-induced platelet aggregation. The investigators of that study5 cautioned against overinterpretation of in vitro alterations in platelet function.
In the present study, the ADP and AA pathways for platelet function were evaluated by use of a modified thromboelastography assay. This assay was developed to evaluate the effect of ADP and AA on platelet function, which could provide information regarding the feasibility of the use of ADP or AA antagonists, such as clopidogrel and acetylsalicylic acid, to treat platelet-based hypercoagulation disorders. The purpose of the present study was to determine whether acepromazine caused clinically relevant inhibition of platelet aggregation, not to identify subtle changes in platelet function. Thus, the concentration of ADP used in the modified thromboelastography assay in the study reported here needed to be sufficient to initiate costimulation and aggregation but not so high as to cause aggregation despite inhibition of the ADP receptor.25 The modified thromboelastography assay was validated for use with human blood samples in a study18 that indicated that results obtained via ADP-activated and AA-activated thromboelastography correlated well with results obtained via optical platelet aggregation. However, the percentage of platelet inhibition determined via the modified thromboelastography assay was subjectively greater than that determined by use of optical platelet aggregation for ADP concentrations < 2μM, which implied that stimulation may have been inadequate. In another study,20 results for a modified thromboelastography assay were similar to results obtained by use of whole blood impedance aggregometry on blood samples from healthy dogs that had been administered clopidogrel. Because the modified thromboelastography assay assesses the strength of the clot, it is possible that subtle changes in initial platelet function or activation could go undetected. Modified thromboelastography allows evaluation of the effects of specific cofactors, such as ADP and TXA2, on platelet activation; however, it cannot differentiate between the effects of specific receptors. To our knowledge, a modified thromboelastography assay has not been used to detect altered platelet function caused by acepromazine administration. It is possible that the platelet agonists used in the present in vitro assay may have overridden any effect caused by acepromazine in vivo. In fact, the use of platelet agonists at concentrations that maximized the MA may have reduced the sensitivity of the thromboelastography assay to detect any subtle effect of acepromazine on platelet function. However, regardless of the concentrations of the platelet agonists used in the modified thromboelastography assay, the results of the present study suggest that if acepromazine does have an effect on platelet function, it is most likely a small effect.
Acepromazine may cause platelet inhibition by mechanisms not evaluated in the present study. Thrombin inhibition is unlikely because the TEGthrombin MA results were not significantly decreased following acepromazine administration. Epinephrine is considered a weak platelet agonist in dogs,26,27 whereas serotonin only induces a change in the shape of platelets but does not induce platelet aggregation or release of granule contents.26,28 Thus, each of these pathways in and of itself is unlikely to contribute to clinically relevant hemorrhage. In studies2,a in which investigators have found changes in platelet aggregation following acepromazine administration, no evidence of clinically relevant hemorrhage or alterations in clotting times or bleeding times were reported. The lack of clinically relevant hemorrhage observed in those studies2,a and the lack of a significant association between acepromazine administration and platelet aggregation detected in the present study may be a result of multiple pathways for platelet activation or the redundancy of platelet activation in vivo.
It is also possible that the modified thromboelastography assay did not adequately assess TXA2-mediated platelet aggregation. An ADP–activated modified thromboelastography assay has been validated in people18 and dogs.20 An AA-activated modified thromboelastography assay has been validated in people18,28 but has not yet been validated in dogs.
In the present study, the doses of acepromazine administered, timing of blood sample collection, and duration of the washout period between treatments may have affected the outcome. The doses of acepromazine administered in this study are common in both clinical and research settings, but administration of higher doses of acepromazine has been reported.2 The use of higher doses of acepromazine could lead to changes in platelet function detectable by use of modified thromboelastography. The plasma half-life of acepromazine is 7.1 hours following IV administration in dogs.29 The purpose of the present study was to evaluate the effect of acepromazine on circulating platelets; thus, the plasma half-life of acepromazine formed the basis for determining the washout period. Because the mechanism by which acepromazine affects platelet function is unknown, it is possible that the mechanism is mediated by some unmeasured metabolite of acepromazine or some effect of redistribution of acepromazine that was not considered. Thus, the washout period may not have been adequate, or acepromazine's effect on platelet function may not have been detectable at our predetermined times for blood sample collection.
Acepromazine can cause a decrease in Hct and platelet count following administration in dogs.30,a A dose of 0.08 mg of acepromazine/kg, IV, caused a decrease in Hct by approximately 15% (eg, Hct decreased from 40% to 33%) in 1 study.30 Changes in Hct have been negatively correlated with MA measurements obtained by use of thromboelastography assaysn; a decrease in Hct will result in an increase in MA, which is indicative of an overall hypercoagulable state. Measurements of MA are particularly affected when Hct is ≤ 20%31 and > 60%,32 which indicates that the correlation between MA and Hct is not linear and supports findings of other investigators.o Mean platelet count in healthy dogs decreased from 247,250 to 185,000 platelets/μL after administration of acepromazine (0.1 mg/kg, IV).a Substantial reductions in platelet count will cause a decrease in MA measurements.33,o In 1 study,o a decrease in platelet count from 300,000 to 100,000 platelets/μL (Hct, 50%) led to a decrease in mean MA from 61 to 50 mm. Investigators33 who used an in vitro model of thrombocytopenia and a rotational thromboelastography assay observed no significant differences in MA measurements until the platelet count was < 65,000 platelets/μL. In the present study, all dogs had Hcts and platelet counts within reference ranges on initial CBCs. A mean total blood volume of < 5% was collected from each dog over the 10-day period during which the study was conducted, and no other interventions were performed. On the basis of the expected changes in hematologic variables following acepromazine administration, the Hcts and platelet counts in the study dogs would not have differed enough to affect the MA measurements obtained with the modified thromboelastography assay. However, Hct and platelet counts were not measured in all blood samples and blood samples were diluted in vitro by the addition of reagents. It is unclear whether these factors had any effect on the results of the present study.
Traumatic or prolonged venipuncture can alter hemostatic activation following blood sample collection.34,35 Vascular injury may lead to activation of coagulation factors and platelets via exposure of subendothelial collagen or tissue factor, and result in an artificially short R, short K, large α angle, or large MA as determined via thromboelastography assays. Excessively traumatic venipuncture has been associated with hypocoagulability caused by depletion of fibrinogen and other clotting factors,34 and difficult venipuncture has been associated with a prolonged R in dogs.p In the present study, only 2 phlebotomies were classified with a difficulty score of 3 (ie, the most difficult); therefore, it is unlikely that traumatic venipuncture had any effect on the MA measurements obtained.
Power calculations for the study reported here indicated that the study had sufficient power to detect clinically relevant changes in MA measurements obtained from the modified thromboelastography assay; therefore, the probability of a type II error was small, and the negative results reported were likely real. However, it is possible that a significant association between acepromazine administration and platelet function would have been found in a study with a larger sample size.
In the present study, no significant changes in platelet function (as determined by use of a modified thromboelastography assay) were detected after a single dose of acepromazine administered IV, and these results differ from those of other studies.2–4,a It is possible that acepromazine causes subtle changes in platelet function that could not be detected with the modified thromboelastography assay used or inhibits platelet function by some other mechanism that was not evaluated. All of the dogs in the present study were healthy, and caution is still warranted when acepromazine is used in patients with suspected or confirmed coagulation disorders. Further studies that assess the effects of acepromazine on hemostasis and platelet function (via pathways other than those evaluated in the present study) in compromised patients are necessary.
ABBREVIATIONS
| AA | Arachidonic acid |
| HDA | High dose of acepromazine maleate |
| LDA | Low dose of acepromazine maleate |
| MA | Maximum amplitude |
| TEGAA | Reptilase-activated thromboelastography with arachidonic acid added |
| TEGADP | Reptilase-activated thromboelastography with ADP added |
| TEGfibrin | Reptilase-activated thromboelastography |
| TEGthrombin | Citrated kaolin–activated thromboelastography |
| TXA2 | Thromboxane A2 |
De Castro LCG, Bechara JN, e Silva MAML, et al. Effects of acepromazine on platelet number and function in dogs (abstr), in Proceedings. Assoc Vet Anaesth Autumn Meet 2007;16.
Haemoscope Corp, Niles, Ill.
Acepromazine maleate, Boehringer Ingelheim Vetmedica Inc, St Joseph, Mo.
BD Vacutainer Safety-Lok blood collection set (Ref 367281), Becton, Dickinson and Co, Franklin Lakes, NJ.
BD Vacutainer 3.0-mL serum blood collection tube (Ref 366668), Becton, Dickinson and Co, Franklin Lakes, NJ.
BD Vacutainer 1.8-mL buffered sodium citrate 3.2% (Ref 366392), Becton, Dickinson and Co, Franklin Lakes, NJ.
BD Vacutainer 4.0-mL lithium heparin 75 USP Units (Ref 367884), Becton, Dickinson and Co, Franklin Lakes, NJ.
TEG 5000 thromboelastograph hemostasis analyzer, Haemoscope Corp, Niles, Ill.
Platelet Mapping Assay Full Assay Kit, Haemoscope Corp, Niles, Ill.
Kaolin, Haemoscope Corp, Niles, Ill.
Calcium chloride 0.2M, Haemoscope Corp, Niles, Ill.
Walker JM, Hanel RM, Hansen BD. Comparison of venous sampling methods for thromboelastography in clinically normal dogs (abstr), in Proceedings. Int Vet Emerg Crit Care Symp 2010;A7.
STATA, version 11, StataCorp, College Station, Tex.
Vilar P, Hansell J, Westendorf N, et al. Effects of hematocrit on thromboelastography tracings in dogs (abstr). J Vet Intern Med 2008;22:774.
Tuman K, Nayler B, Spiess S, et al. Effects of hematocrit on thromboelastography and Sonoclot analysis (abstr). Anesthesiology 1989;71:A414.
Garcia-Pereira BL, Scott MA, Koenigshof AM, et al. Effect of venipuncture quality on thromboelastography in healthy dogs (abstr), in Proceedings. Int Vet Emerg Crit Care Symp 2010;A3.
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