Haloperidol affects coupling between QT and RR intervals in guinea pig isolated heart. Petr Vesely, Tibor Stracina, Miroslava Hlavacova, Josef Halamek, Jana ...
Haloperidol affects coupling between QT and RR intervals in guinea pig isolated heart Petr Vesely, Tibor Stracina, Miroslava Hlavacova, Josef Halamek, Jana Kolarova, Veronika Olejnickova, Veronika Mrkvicova, Hana Paulova, Marie Novakova
Supplementary Materials to Manuscript
Guinea Pig Isolated Heart as a Model for the studying of haloperidol effect on the QT/RR coupling Guinea pig isolated heart is eligible model for studying of direct drug effects on heart activity. Guinea pig as a model organism was used in this study, since guinea pig cardiac cells exhibit specific ion channels quite comparable to those of humans.1 Haloperidol was administered via intraperitoneal route in the dose identical to our previous studies.2,3 Pharmacokinetics of intraperitoneal administration is comparable to oral administration because the primary route of absorption is into portal vein system and passage through liver.4,5 It is similar to clinical situation since the majority of long-term haloperidol-treated patients use the oral form.6 For the studying of the relationship between QT and RR intervals, spontaneously beating isolated heart model perfused according to Langendorff was used. Langendorff isolated heart is eligible model for studying of direct drug effects on heart activity without influence of extracardial systems (such as endocrine and autonomic nervous regulation).7 Noticeable advantage of the isolated heart model is easy way how to administer strictly defined concentrations of drug directly to coronary system of the heart and may simulate an acute i.v. application in clinical practice. Concentration of haloperidol applied in the experiment is relevant to reported plasma levels in patients treated with haloperidol.8 Since the heart rate of spontaneously beating isolated heart is affected by even small changes of experimental conditions (such as temperature and perfusion pressure), experiments were done in strictly
controlled environment and precise monitoring of experimental conditions was applied. Some of the isolated heart model limitations (in short-lasting experiment) are certain influence of ischemia to which is the heart exposed during preparation and the fact that direct effects of the drug on ionic currents cannot be studied. One of the model limitations is certain influence of ischemia to which the heart is exposed during preparation.9 Preparation ischemia increases the production of reactive oxygen species in the heart tissue. Moreover, haloperidol increases oxidative stress in various tissues.10-12 During increased oxidative stress, HNE is produced by peroxidation of membrane lipids.13 To evaluate the quality of the isolated heart preparation in our study and to detect possible influence of preparation ischemia, assessment of CK, LD, lactate, and HNE was performed. In our experimental set-up, the biochemical examination was limited by lack of coronary effluent sample at the end of stabilization period as well as at the end of haloperidol exposures. These samples could not be obtained due to need of continuous electrogram recording. However, based on the above findings, we may conclude that the preparation ischemia in our experimental set-up does not affect the reliability of the obtained results.
Dose of Haloperidol and its Administration Doses of haloperidol for both chronic and acute administration were chosen according to the previous studies.2,3 For chronic administration, haloperidol (Sigma Aldrich, USA) in dose of 2 mg/kg or vehiculum (2% ethanol in aqua pro injectione) in identical volume were administered by intraperitoneal injection once a day for 21 consecutive days. The dose of haloperidol or vehiculum was calculated daily for each animal according to its actual body mass. Weighing and application was done always at the same daytime (around noon). No anaesthetic or analgesic agents were
used. Gentle handling and quiet approach were applied to reduce discomfort of animals during the manipulation. For acute administration, haloperidol prediluted in ethanol was administered diluted in KrebsHenseleit solution at the concentration of 10 nM (ethanol in concentration of 8.22 nM in the final solution). The Krebs-Henseleit solution with haloperidol was administered into isolated
QT correction QT was corrected to the heart rate according to subject specific linear correction model based on QT/RR coupling.14 Such method is more appropriate than corrections according to Bazett or Fridericia.15 In phases H1, W1, H2, and W2, the QTc was calculated according to formula:
QTc= a1*RR0+a0
where a1 and a0 are parameters of individual QT/RR coupling in particular experimental phase, and RR0 is a nominal RR interval. For guinea pig isolated heart, RR0 was set to 400 ms. In stabilisation phase, non-linear changes of QT/RR coupling should be expected, therefore it is not possible to apply the linear model. Non-linearity in QT/RR coupling during stabilisation relates to adaptation of isolated heart to experimental conditions. The isolated heart should be considered as stable (and therefore linear character of QR/RR coupling might be expected) in short period at the end of stabilisation phase. However, the stable period (not more than 2 minutes of the record) is too short for accurate calculation of QT/RR coupling parameters. Therefore, modified subject specific linear correction model was applied. The QTc was calculated according to formula:
QTc= QT+a1*(RR-RR0)
where the RR0 is nominal RR interval (RR0=400 ms). The parameter a1 was calculated as the average value of the QT/RR coupling computed over all phases of the experiment except of the stabilization.
Figure S1: An example of measured (Pperf and Tperf) and detected (RR and QT) signals in isolated heart experiment. The vertical lines show the onset of individual phases of the experiment. Pperf – perfusion pressure, Tperf – temperature of perfusion solution
Table S1: The levels of creatine kinase, lactate dehydrogenase and lactate in the samples of coronary effluent. C
CK
T
sample 1
sample 2
2.32
0.00
P
sample 1
sample 2
1.25
0.00
0.012 -1
[nkat.min ]
(1.80; 4.90)
(0.00; 0.87)
LD
37.11
22.93
0.028 (0.82; 2.50)
(0.00; 0.34)
32.49
19.98
0.017 -1
[nkat.min ]
(32.23; 39.78)
(20.70; 25.33)
lactate
2.02
0.00
0.046 (26.68; 35.30)
(16.94; 24.46)
1.44
0.48
(0.96; 2.54)
(0.00; 1.27)
0.028
[μmol.min ] -1
(0,70; 2.93)
(0.00; 1.27)
P
0.028
Values are expressed as median (25th; 75th quartile). C – control group, T – haloperidol treated group, sample 1 – at the end of the first minute of stabilization (S) phase, sample 2 – at the end of experiment (last minute of W2 phase), P – P value (P < 0.05 indicates significant difference), CK – creatine kinase, LD – lactate dehydrogenase
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