Results
Alfaxalone pharmacodynamics for Lewis rats
Arterial blood pressure measurements for the Lewis rats during
anaesthesia are presented in Figure 2. All rats showed an initial
short-lived decrease in mean arterial blood pressure (MAP), heart rate
and respiratory rate because of concomitant administration of isoflurane
and alfaxalone. Within 5 minutes following discontinuation of
isoflurane, all rats demonstrated an increase in blood pressure from the
baseline readings obtained during anaesthesia with the gaseous volatile
agent. Blood pressure (mean, systolic, diastolic), heart rate and
respiratory rate at baseline were not significantly different between
male and female rats under isoflurane anaesthesia. Heart rates remained
stable during alfaxalone anaesthesia and there was no significant
difference between the sexes at any time points. Systolic, mean and
diastolic arterial pressures all increased from baseline under
isoflurane anaesthesia. The MAP remained elevated until 220 minutes for
males before declining, while females showed early signs of a MAP
decrease from between 40-75 minutes. Significant differences in MAP were
detected between the sexes at 120 minutes (p < 0.05).
All Lewis rats required IPPV as judged by apnoea, or a rise in end tidal
carbon dioxide coupled with a decrease in respiratory rate and effort at
some point and for different time periods. Blood gas parameters and
biochemistry values are presented in Table 1. There were no significant
differences between sexes for all parameters.
Hypnotic effect
The plane of anaesthesia was continually evaluated by serial
cardiopulmonary measurements, blood gas analysis and reflex responses.
Subjective evaluation of this hypnotic effect of the alfaxalone in all
Lewis rats was excellent.
Deterministic individual alfaxalone PK for Lewis rats
A 1-compartment infusion model was shown to have the best fit to the
individual Lewis rat PK data according to AIC. Figure 3 compares the
male versus female Lewis rat PK data and model fit (curves) for each
rat. Plasma concentrations of alfaxalone during maintenance CRI were
greater in the females compared to male rats. PK parameters obtained
from the model fit are shown in Table 2. Logarithmic transformed
pharmacokinetic parameters were shown to be significantly different
between the male and female rats. Mean CL of alfaxalone for male rats
was more than twice that of female rats and with a higher Vdss for the
latter, resulted in an almost 5 fold longer half-life in female rats
compared to males.
NLME PK model for alfaxalone (population 1)
The most parsimonious NLME model obtained for population 1 was a
1-compartment model with random effects included on all parameters and
no correlation (diagonal omega matrix). Goodness of model fit can be
found in the supplementary data file. The covariates for sex and rat
strain had the most significant influence on clearance (CL) and rat
strain having the most significant influence on volume of distribution
(Vd). The CL and Vd for individual rats (expressed per total body
weight) within the population model are described as follows:
CL = CLTV * e(-0.841 * sex covariate)* e(0.478 * strain covariate) * e(CL
eta) Equation 1
Vd = VdTV * e(-0.0237 * strain
covariate) * e(Vd eta) Equation 2
Where CLTV and VdTV are the typical
values (fixed effect) for Clearance (25.2 ml min-1)
and volume of distribution (0.57 L) within population 1. These fixed
effects are adjusted by the covariates (0 for male and Lewis, 1 for
female and Sprague Dawley) to give an adjusted typical value for each
group. CL eta and Vd eta represent the random effects, such as
inter-individual variability, in the population for clearance and volume
of distribution. Sex and strain outputted adjusted typical values (per
total bodyweight) and post hoc PK parameters (per kg bodyweight) for the
most parsimonious NMLE model are shown in Table 3. These PK parameters
were encompassed by the 2.5 and 97.5% confidence intervals of the
bootstrap resampling analysis indicating a robust model. Mean arterial
pressure plotted against plasma alphaxalone concentration for male Lewis
and female Lewis rats is depicted in Figure 4 (A and B). However, MAP
decreases for female Lewis rats (B) were evident when alfaxalone
concentration exceeded approximately 20 µg ml-1.
Adjusted alfaxalone infusion regimen for female SD rats
The NLME PK model for alfaxalone using population 1 was used to design
an adjusted alfaxalone infusion regimen for female SD rats that matches
male SD plasma concentrations minimising cardiopulmonary depression but
maintaining an appropriate hypnotic effect.
Alfaxalone pharmacodynamics for SD male and females
There was no statistical difference between dose adjusted female and
male SD rats mean arterial blood pressure (Figure 5).
IPPV was required for 9/16 female SD, and 5/8 male SD rats. Blood gas
parameters and biochemistry values are presented in Table 1. There were
no significant differences between sexes or strains for all parameters.
Hypnotic effect
The plane of anaesthesia was deemed inadequate for injection of
capsaicin as part of the antinociception study for two female SD rats
during the final reduced phase of the CRI in view of a very faint
sluggish paw withdrawal and spontaneous blinking. No rats demonstrated
gross purposeful movement or required a change in the infusion rate to
improve the plane of anaesthesia however surgical anaesthesia was not an
outcome measure of the model.
Alfaxalone pharmacokinetics for SD males and females
Figure 6 shows the measured alfaxalone plasma concentrations versus time
for male and female SD rats using the adjusted regimen for the latter
along with the simulated median, upper and lower 95% confidence
intervals using the NMLE PK model. Plasma concentrations were similar
with no statistical difference between male and female SD rats during
the plateau phase.
Figure 7 shows no relationship between MAP and alfaxalone concentration
for male SD (A) and dose adjusted female SD (B) rats.
NLME pharmacokinetic model for alfaxalone (population 2)
The most parsimonious NLME model obtained for population 2 was again a
1-compartment model with random effects included on all parameters and
no correlation (diagonal omega matrix). Goodness of model fit can be
found in the supplementary data file. The covariates for LCBW and sex
had the most significant influence on alfaxalone clearance. Similar to
the model for population 1, rat strain had the most significant
influence on Vd. The CL and Vd for individual rats (expressed for total
body weight) within the population model are described as follows:
CL = CLTV * (1 + LCBW * 3.64) * e(-0.43
* sex covariate) * e(CL eta) Equation 3
Vd = VdTV * e(-0.692 * strain
covariate) * e(Vd eta) Equation 4
where CLTV and VdTV are the typical
values for Clearance (35.2 ml min-1) and volume of
distribution (0.51 L) within population 2. These fixed effects are
adjusted by the covariates (0 for male and Lewis, 1 for female and
Sprague Dawley and LCBW) to give an adjusted typical value for each
group. Sex and strain outputted adjusted typical values and post hoc PK
parameters (normalised per kg bodyweight) for the most parsimonious NMLE
model are shown in Table 4. These PK parameters were encompassed by the
2.5 and 97.5% confidence intervals of the bootstrap resampling analysis
indicating a robust model.