Department of Pharmaceutics, Faculty of Pharmaceutical
Sciences, The University of Tokyo, Bunkyo-ku, Tokyo 113, Japan
A distributed model has been used to clarify the mechanism of the
restricted and differential distribution of the quinolone antibiotics
in the rat central nervous system (CNS). The symmetrical permeability
clearances across the blood-brain barrier (BBB), PSBBB, and
across the blood-cerebrospinal fluid barrier (BCSFB), PSCSF, and the active efflux clearances across the BBB,
PSBBB,eff, were obtained from a nonlinear least squares
regression analysis combined with the fast inverse Laplace transforming
program for in vivo data. The values of
PSBBB,eff were 10- to 260-fold greater than those of
PSBBB, providing kinetic evidence to support the hypothesis
that a significant efflux transport across the BBB is responsible for
the limited distribution of quinolones in brain tissue. Moreover, by
simulation studies, we could demonstrate the concentration profiles in
the brain as a function of the distance from the ependymal surface.
However, active efflux transport across the BCSFB has been suggested to
have only a slight effect on the apparent elimination from the
cerebrospinal fluid. Comparing the apparent brain tissue-to-unbound
serum concentration ratio at steady state, it has been suggested that
the net flux across the BBB, i.e., the ratio of
PSBBB to the sum of PSBBB and
PSBBB,eff, is a determinant for the differential
distribution of these quinolones in brain tissue. Such a putative
active efflux transport system would play a significant role in
decreasing the brain interstitial fluid concentration of quinolones.
 |
Introduction |
The
side effect of quinolone antimicrobial agents (quinolones) on the CNS
such as confusion, hallucinations, anxiety, agitation, depression and
convulsive seizures is one of the most serious problems associated with
their use as chemotherapeutic agents (Christ, 1990
). Because the
interaction of quinolones with the receptor of GABA in the brain is
responsible for CNS side effects (Akahane et al., 1989), it
is important to understand the mechanism for the distribution of
quinolones in brain tissue and the CSF in quantitative terms. Several
investigators have reported that quinolone concentrations in brain
tissue and CSF are lower than in serum after systemic administration
(Ichikawa et al., 1992; Sato et al., 1988). In
addition, we previously demonstrated the presence of such an active
transport from the CSF to blood across the BCSFB by examining the
efflux of quinolones from the CSF after intracerebroventricular
administration (Ooie et al., 1996b) and by examining the
uptake of quinolones by the isolated choroid plexus (Ooie et
al., 1996a). We have also demonstrated that brain-to-plasma and
CSF-to-plasma unbound concentration of quinolones are less than unity
at steady state (Ooie et al., 1996c). However, no report has
appeared in which the efflux transport for quinolones across the BBB
has been characterized. To clarify the mechanism for the restricted
distribution in the brain tissue and the CSF, one of the best
ways is to apply pharmacokinetic model analysis as reported previously
(Dykstra et al., 1993; Ogawa et al.,
1994; Sato et al., 1988; Wang and Sawchuk, 1995).
Considering the anatomical features of the brain tissue and the CSF,
the distributed model (Collins and Dedrick, 1983; Suzuki et
al., 1997) will provide much more information about the kinetics
of drug diffusion through the brain tissue.
The present study investigated the process governing the restricted
distribution of quinolones in the CNS based on the distributed model.
In this study, we determined the initial CNS uptake of quinolones in
rats. Moreover, these data, along with previously published data
[brain-to-plasma and CSF-to-plasma concentration at steady state (Ooie
et al., 1996c) and CSF concentration profiles after
intracerebroventricular administration (Ooie et al.,
1996b)] were kinetically analyzed to establish the presence of active transport for the efflux of quinolones from the brain to blood across
the BBB. To clarify the mechanism of the differential distribution of
quinolones in the CNS, a comparative kinetic analysis was also performed for six quinolone analogs: NFLX, AM-1155, OFLX, FLRX, SPFX
and PFLX.
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Materials and Methods |
Materials.
All the quinolones, NFLX, AM-1155, FLRX, OFLX,
SPFX and PFLX were synthesized in the Central Research Laboratories of
Kyorin Pharmaceutical Co., Ltd. (Tochigi, Japan). All other chemicals were commercial products of analytical grade. Male Wistar rats weighing
250 to 300 g (Japan Laboratory Animals, Inc., Tokyo, Japan) were
used throughout the experiments, which were conducted according to the
guidelines provided by the Institutional Animal Care Committee (Faculty
of Pharmaceutical Sciences, The University of Tokyo).
In vivo distribution of quinolones.
Rats were
anesthetized with an i.p. dose of 1.5 g/kg ethylcarbamate, and
cannulation with polyethylene tubing (PE-50) was performed into the
femoral artery and vein. An i.v. bolus dose of quinolone (10 mg/kg) was
administered through the femoral vein cannula. At fixed times after
injection of each quinolone, arterial blood specimens were withdrawn
and centrifuged to obtain serum. At 1, 3, 5 and 10 min after i.v.
administration, crystal-clear CSF specimens were obtained from
individual rats by cisternal puncture with a 24-gauge needle
(Matsushita et al., 1991). Immediately after collection of
the CSF, each rat was decapitated and the cerebral cortex removed.
Determination of drug concentrations.
Brain tissues were
homogenized with 4 volumes of 0.067 M phosphate buffer (pH 7.0). After
centrifugation of tissue homogenates, supernatants were used to measure
drug concentrations. The concentration of quinolones in serum, tissue
homogenates and CSF was determined by the HPLC as described previously
(Ooie et al., 1997).
Kinetic analysis of serum concentration-time profiles.
With
the previously reported values for the serum unbound fraction (Ooie
et al., 1996c), the unbound serum concentration of each
quinolone (Cp,u) was obtained from the
in vivo study after i.v. administration. The area under the
unbound serum concentration-time curve for time t
(AUCu,0-t) of each quinolone was estimated by the linear trapezoidal rule. The apparent influx clearance across
the BBB and BCSFB (PSBBB,app and
PSCSF,app) were obtained from in vivo
data by dividing the brain or the CSF concentration at 1 min by the
corresponding AUCu,0-1 min after i.v.
administration; these were used as initial estimates for the
distributed model analysis described as follows. Mean
Cp,u values obtained from 6 to 12 rats
after i.v. administration of each quinolone versus time
profiles were examined by nonlinear least squares regression analysis
(Yamaoka et al., 1981) by the following equation:
|
(1)
|
Distributed model analysis.
A distributed model (Collins and
Dedrick, 1983; Suzuki et al., 1997) has been used to analyze
quinolone distribution in the CNS. The analysis was performed according
to the method described previously (Suzuki et al., 1997)
with some modifications. A diagrammatic representation of an anatomical
compartment in the brain tissue and the CSF is presented in figure
1. In estimating the brain tissue
concentration-time or the CSF concentration-time profiles the following
assumptions were made: 1) The brain tissue and the CSF are described by
a one-dimensional slab of tissue. 2) Drug permeates through the BBB and
the BCSFB in both directions, i.e., influx and efflux. 3)
There is a constant bulk flow of the CSF. 4) Drug concentration in
brain ISF at the ependymal surface is the same as that in the CSF. 5)
Drug diffuses through the brain tissue according to Fick's law of
diffusion. 6) Drug distributes into the intracellular fluid space in
the brain. Based on this model (fig. 1), the Laplace transformed
equations of total brain (Cbr) and CSF
concentration (CCSF) after i.v. or
intracerebroventricular administration were obtained.
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Fig. 1.
Distributed model for the kinetic analysis of
quinolone distribution between brain tissue, CSF and blood.
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Equation 2 represents a mass balance equation describing the
drug concentration in the brain tissue:
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(2)
|
where Cbr(x,t)
is the drug concentration in the brain tissue at a distance,
x, from the ependymal surface at time t.
Dt, PSBBB and
PSBBB,eff represent the apparent diffusion
coefficient through the brain tissue, the permeability clearance of
symmetrical transport across the BBB and the permeability clearance of
the active efflux process from the brain ISF to circulating blood across the BBB, respectively.
Cp,u(t) is the unbound drug
concentration in the serum at time t,
Vbr is the distribution volume in the brain
tissue defined as the concentration ratio of total brain-to-ISF.
Equation 3 represents a mass balance equation describing the drug
concentration in the CSF:
|
(3)
|
where CCSF(t) is the drug
concentration in the CSF at time t,
VCSF is volume of the CSF,
PSCSF is the symmetrical permeability clearance
across the BCSFB, PSCSF,eff is the active efflux
clearance across the BCSFB, Q is the bulk flow
rate of the CSF and Ar is the surface area of the cerebroventricular
ependyma. Assuming that the ISF concentration
(CISF) at the ependymal surface is the same
as the CSF concentration, i.e.,
CCSF(t) = CISF(0,t) = Cbr(0,t)/Vbr,
equation 3 can be converted to equation 4 as a boundary condition of
equation 2, at x = 0.
|
(4)
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At some distance (x*) from the ependymal surface, CSF
events no longer create a driving force for diffusion flux, so that other boundary conditions of equation 2 can be defined, at
x 
x*:
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(5)
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Moreover, there is no drug in brain tissue and the CSF at time
t = 0. Thus, we have used the following relationship as
an initial condition of equations 2 to 5:
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(6)
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Taking the Laplace transform of equations 2 to 5, equations 7
and 8 can be obtained for the drug concentration in brain and brain
ISF, respectively, at a distance x:
|
(7)
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and
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(8)
|
where s is the operator of time t. In this
calculation, we assumed that
Cp,u(t) is given by equation 1
and x* 
. Defining the thickness of cerebral cortex
surrounding the CSF as L, the following equation
can be derived for the average drug concentration in the
brain,
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(9)
|
Moreover, considering the condition that x
is zero, i.e., the CSF concentration at time t,
the following equation can be obtained from equations 7 and 8;
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(10)
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With equations 9 and 10, the area under
concentration-time curve of average brain tissue and the CSF,
i.e., AUCbriv and AUCCSFiv,
respectively, can be obtained as:
|
(11)
|
and
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(12)
|
With use of equations 11 and 12, the brain-to-unbound serum
concentration ratio (Kp,u,br) at steady
state and the CSF-to-unbound serum concentration ratio
(Kp,u,CSF) at steady state can be obtained as the ratio of AUCbriv to the area under the unbound
serum concentration-time curve (AUCu) and that of
AUCCSFiv to AUCu by the following
equations:
|
(13)
|
and
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(14)
|
For an intracerebroventricular administration, mass balance
equations describing the drug concentration in the brain tissue and the
CSF can be obtained as equations 15 and 16, respectively.
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(15)
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(16)
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Assuming that
CCSF(t) = CISF(0,t) = Cbr(0,t)/Vbr,
the following equa tion can be obtained as a boundary condition of
equation 15, at x =0:
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(17)
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With the boundary condition that a distance x is
significantly greater than x*, the following relation is
obtained, at x x*.
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(18)
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As an initial condition of equation 15, the following
relationship is obtained for the CSF concentration at time zero:
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(19)
|
Taking the Laplace transform of equations 15 to
17, the following equations can be obtained.
|
(20)
|
and
|
(21)
|
With use of equation 21, the area under concentration-time
curve of the CSF after an intracerebroventricular administration (AUCCSFicv) was obtained as:
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(22)
|
Accordingly, the efflux clearance from the CSF
after an intracerebroventricular bolus administration
(CLCSF) is described by the following equation.
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(23)
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Estimation of BBB and BCSFB permeability.
For the model
analysis, the fixed parameters were obtained in the following way.
Regarding Dt of quinolones, the diffusion coefficient of quinolones in agar (Dw), was
estimated from a previous report based on the molecular weight
(Fenstermacher and Kaye, 1988). Based on the previously reported
relationship (Fenstermacher and Kaye, 1988) between the ratio of
Dt to Dw and
Vbr, defined as the ratio of the
Cbr and CISF,
the Dt value of each quinolone was
estimated and is listed in table 1. The
Vbr values of NFLX, OFLX, FLRX and PFLX
were taken from a previous report which was determined by using a brain
microdialysis (Ooie et al., 1997). For AM-1155 and SPFX, the
Vbr value was estimated from the mean Vbr value of NFLX, OFLX, FLRX and PFLX,
i.e., 1.94 ± 0.36 ml/g brain (mean ± S.E.).
PSCSF,eff was obtained from the uptake study using the isolated choroid plexus reported previously (Ooie et al., 1996c). The serum unbound fraction (fu)
of quinolones, Q, VCSF, Ar, and
L were taken from previous reports (Cserr and Dyke 1971;
Ogawa et al., 1994; Ooie et al., 1996c; Suzuki
et al., 1985) and are listed in tables 1 and
2. The total brain
concentration-to-unbound serum concentration ratio
(Kp,u,br) at steady state and the
CSF-to-unbound serum concentration ratio
(Kp,u,CSF) at steady state after i.v. constant infusion, and CLCSF were taken from
previous reports (Ooie et al., 1996b,c) and are listed in
table 1.
By use of equations 9, 10 and 21, PSBBB,
PSBBB,eff and PSCSF were
determined simultaneously by a nonlinear least squares regression analysis program combined with the fast inverse Laplace transform algorithm (MULTI(FILT)) developed previously (Yano et
al., 1989). For the model fitting, the following in
vivo data were used simultaneously: 1)
Cbr-time profile after i.v. bolus
administration (data shown in fig. 2B as an integration plot); 2)
CCSF-time profile after i.v. bolus
administration (data shown in fig. 2C as an integration plot); 3) The
value of Kp,u,br at steady state after a
constant i.v. infusion (table 1); 4) The value of
Kp,u,CSF at steady state after constant
i.v. infusion (table 1); 5) CCSF-time
profile after intracerebroventricular bolus administration (taken from a previous study; Ooie et al., 1996b).
Cp,u-time profiles of quinolones descried
above were substituted for A, B, 
and 
in equations 9
and 10. The initial values of PSBBB and
PSCSF were set as the apparent influx clearance
obtained in in vivo experiment,
PSBBB,app and PSCSF,app,
respectively. The initial value of PSBBB,eff was assumed to be zero. The PSBBB and
PSCSF were allowed to vary within a range of
±50% of its initial value.
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Fig. 2.
Unbound serum concentration-time profile (A), total
brain-to-unbound serum concentration ratio
(Kp,u,br; B) versus the ratio of the area under the unbound serum concentration time curve
(AUCu) to the unbound serum concentration
(Cp,u), and CSF-to-unbound serum
concentration ratio (Kp,u,CSF; C)
versus the ratio of the area under the unbound serum
concentration time curve (AUCu) to the unbound serum
concentration (Cp,u) of quinolones in rats. Six different quinolones (10 mg/kg) were administered intravenously and
blood samples were collected. At 1, 3, 5 and 10 min after dosing, CSF
was obtained from individual rats by cisternal puncture. Immediately
after the collection of CSF, the cerebrum was removed. Each point
represents the mean ± S.E. of 4 to 12 animals. Where error bars
are not shown, the S.E. is contained within the symbol.  , NFLX;  ,
AM-1155;  , FLRX;  , OFLX;  , SPFX;  , PFLX.
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The concentration of quinolone in brain tissue was determined by
subtracting the quinolone concentration in the brain vascular space
from the observed total brain concentration. The blood vascular volume
was assumed to be 0.020 ml/g from the reported plasma vascular volume
(0.011 ml/g; Pardridge et al., 1991) using a hematocrit of
0.45. This value has been reported as the distribution space of mouse
immunoglobulin G in rat brain (Pardridge et al., 1991). Reed
and Woodbury (1963) obtained a value of 0.01 ml/g for the distribution
of [14C]inulin (5,000 Da) in rat brain, whereas
they obtained value of 0.005 ml/g for
[131I]serum albumin (69,000 Da). With use of
sucrose, Smith et al. (1988) also reported that the vascular
space in rat brain is approximately 0.007 ml/g brain. Because we did
not measure the cerebral vascular space in each rat, variations in this
value may affect the analysis, in particular for quinolones (such as
NFLX) whose brain distribution is limited.
Simulation of drug concentration in the CNS.
The equations
for Kp,u,br (equation 13) and
Kp,u,CSF (equation 14) at steady state
after i.v. administration and for CLCSF after intracerebroventricular administration (equation 23) were used in the
simulation study. To predict the CISF in
various regions of the CNS, equations 8 and 20 were used. Unbound serum
concentration versus time profiles of three different
quinolones (NFLX, FLRX and PFLX) were taken from previous reports
(Ichikawa et al., 1992; Jaehde et al., 1992;
Kusajima et al., 1986) and were used for the estimation of
CISF after i.v. bolus administration at a
dose of 10 mg/kg. Furthermore, CISF after
intracerebroventricular administration at a dose of 10 µg/animal was
predicted with equation 20. For the simulation study, Laplace
transformed equations (equations 8 and 20) were analyzed using the fast
inverse Laplace transform program (FILT; Yano et al., 1989).
Data analysis.
Model calculation and fitting were carried
out on an IBM RISC System/6000 work station using AIX XL FORTRAN
Compiler/6000. The results of kinetic analysis are expressed as
means ± calculated S.D. except when noted otherwise. Statistical
analysis was performed by Student's t-test. Correlation was
tested by Pearson's product moment correlation coefficient (r).
| |
Results |
CNS distribution of quinolones after i.v. bolus
administration.
The concentrations in the serum, brain tissue and
the CSF were determined after i.v. administration of 10 mg/kg of each
quinolone. By using the previously reported fu
for each quinolone (table 1; Ooie et al., 1996c), the
Cp,u-time profiles were obtained and are
shown in figure 2A. The pharmacokinetic
parameters to describe the curve were obtained by nonlinear least
squares regression analysis and are listed in table 1. Figure 2B
illustrates initial uptake of quinolones into the CNS after i.v.
administration as an integration plot
(Kp,u,br vs.
AUCu/Cp,u; Patlak et al.,
1983). In principle, extrapolation of the
Kp,u,br versus
AUCu/Cp,u line yields
the cerebral vascular volume as the y-intercept. As shown in
figure 2B, however, the values of the y-intercept for some quinolones are much higher than the vascular space; e.g.,
0.13 and 0.14 ml/g brain for SPFX and PFLX, respectively. These results may be accounted for by considering the rapid passage of these two
quinolones across the BBB. To quantify this rapid passage by model
analysis, the PSBBB,app value was determined with
the early time point (1 min after i.v. administration) data, rather than the slope of the integration plot, assuming no adsorption of
quinolone to the luminal surface of brain capillaries and no back flux
from the brain into the circulating blood. The
PSBBB,app values for six kinds of quinolone
antibiotics (table 1) were further used as the initial value for
PSBBB in the kinetic analysis. Comparing the
values of PSBBB,app, NFLX and PFLX were the
smallest and greatest, respectively, and a 35-fold difference was
observed between them. Figure 2C illustrates the
Kp,u,CSF versus AUC
u /Cp,u plot. Similarly,
PSCSF,app values were determined and are listed
in table 1. Comparing the values of PSCSF,app,
NFLX and SPFX were the smallest and greatest, respectively, and a
27-fold difference was observed between them.
Distributed model analysis.
By use of equations 9, 10 and 21,
PSBBB, PSBBB,eff and
PSCSF were obtained by nonlinear least squares
regression analysis. Figure 3 represents
the comparison between observed and fitted values for the six
quinolones. As shown in figure 3, A, C, D and E, a fairly good
coincidence was observed between observed and model-fitted values for
the brain concentration after i.v. bolus administration, the steady
state Kp,u,br and
Kp,u,CSF after i.v. constant infusion and
the CSF concentration after intracerebroventricular bolus administration. For the CSF concentration after i.v. bolus
administration, the model values were relatively smaller than the
observed values (fig. 3B).
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Fig. 3.
Comparison of fitted and observed values for
quinolone distribution in the CNS. (A) total brain concentration after
i.v. bolus administration; (B) CSF concentration after i.v. bolus
administration; (C) brain-to-unbound serum concentration ratio
(Kp,u,br) at steady state, the observed
values were taken from a previous report (Ooie et al.,
1996c); (D) CSF-to-unbound serum concentration ratio
(Kp,u,CSF) at steady state, the observed
values were taken from a previous report (Ooie et
al., 1996c); (E) CSF concentration after
intracerebroventricular administration, the observed values were taken
from a previous report (Ooie et al., 1996b). , NFLX;
, AM-1155; , FLRX; , OFLX; , SPFX; , PFLX.
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The fitted parameters for the quinolones (PSBBB,
PSBBB,eff and PSCSF) are
summarized in table 3. The
PSBBB values of SPFX and PFLX were about 80-fold
greater than the PSBBB value of NFLX. Moreover,
the PSBBB,eff value was 11- to 260-fold greater
than the PSBBB value (table 3). With use of the
fitted parameters obtained, the net flux across the BBB
[PSBBB/(PSBBB + PSBBB,eff)] was also calculated. A good
correlation was observed (r = 0.93, P < .01) between net
flux (table 3) and steady state Kp,br
values (table 1). PSCSF of NFLX was smaller than
Q, whereas the PSCSF of SPFX was
5-fold greater than Q (table 3). Assuming that
PSBBB,eff equals zero, the
Kp,u,br values were simulated to be
approximately 5- to 10-fold greater than the observed values (data not
shown).
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TABLE 3
Permeability clearance of quinolones obtained by nonlinear least
squares regression analysis combined with a fast inverse Laplace
transform algorithm (MULTI(FILT))
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As shown in figure 4A, a fairly good
correlation was observed between the PSBBB and
the octanol-water partition coefficient (Ooie et al., 1996c)
for the quinolones examined (r = 0.90, P < .01). There was
also a good correlation between the PSCSF and the
octanol-water partition coefficient (r = 0.88, P < .01), but not for the PSBBB,eff (r = 0.52, not
significant).
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Fig. 4.
Relationship between the octanol-water partition
coefficient (log Papp) and symmetrical permeability
clearance across the BBB (PSBBB; A), symmetrical
permeability clearance across the BCSFB (PSCSF; B) and
asymmetric clearance across the BBB (PSBBB,eff; C). The
straight line represents the result of linear regression analysis. ,
NFLX; , AM-1155; , FLRX; , OFLX; , SPFX; , PFLX.
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Because the penetration of ligands across the erythrocyte membrane is
rapid enough (Simanjuntak et al., 1991), the unbound plasma
concentration equals the unbound blood concentration by their
definition. The kinetic parameters defined for the plasma unbound
concentration, therefore, can be used if these values are defined for
the unbound blood concentration.
Prediction of brain ISF concentration as a function of the
distances from the ependymal surface.
Figure
5 represents the
CISF-time profiles as a function of the
distance from the ependymal surface after i.v. bolus administration of
NFLX, FLRX and PFLX at a dose of 10 mg/kg to rats. A significant gradient of CISF was observed as a function
of the distance from the surface. No significant difference in
CISF was observed at cerebral regions more
than 1 mm distant from the ependymal surface (fig. 5).
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Fig. 5.
Prediction of concentration-time profile of
quinolone distribution in various regions of the CNS and serum after
i.v. bolus administration of quinolones, 10 mg/kg, to rats. (A) NFLX;
(B) FLRX; (C) PFLX. Values close to the line represent the distance from the ependymal surface.
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Figure 6 represents the
CISF-time profiles as a function of the
distance from the ependymal surface after intracerebroventricular bolus
administration of NFLX, FLRX and PFLX at a dose of 10 µg/animal to
rats. During the terminal phase, an approximately 10- and 1000-fold difference was observed between the CSF and ISF concentrations at a
distance of 0.5 mm and 1 mm from the surface of ependymal cell layer,
respectively (fig. 6).
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Fig. 6.
Prediction of brain ISF concentration-time profiles
in various regions of the CNS after intracerebroventricular
administration of quinolones, 10 µg/animal, to rats. (A) NFLX; (B)
FLRX; (C) PFLX. Values close to the line represent the distance from
the ependymal surface.
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Effect of diffusion through brain tissue and active efflux
clearance across the BCSFB on Kp,u,br and
Kp,u,CSF at steady state.
The
equation representing Kp,u,br and
Kp,u,CSF at steady state were obtained as
equations 13 and 14, respectively. Assuming that the diffusion
coefficient through the brain tissue is zero, the
Kp,u,br and the
Kp,u,CSF values were estimated and compared with the fitted values in figure 7, A and
B, respectively. The simulated Kp,u,br
value was lower than that of the fitted value, whereas the simulated
Kp,u,CSF value was greater than the fitted value for all quinolones studied. Figures
8, A and B, represent the effect of the
active efflux clearance across the BCSFB on the
Kp,u,br and the
Kp,u,CSF values, respectively. Only a
slight difference was observed between the fitted and simulated values even if PSCSF,eff was assumed to be zero.
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Fig. 7.
Comparison between fitted (solid bars) and
simulated (open bars) Kp,u,br and
Kp,u,CSF values assuming no diffusion
through brain parenchyma tissue. (A) brain-to-unbound serum
concentration ratio at steady state; (B) CSF-to-unbound serum
concentration ratio at steady state.
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Fig. 8.
Comparison between fitted (solid bars) and
simulated (open bars) Kp,u,br and
Kp,u,CSF values assuming no active efflux
via a BCSFB (PSCSF,eff = 0). (A)
brain-to-unbound serum concentration ratio at steady state; (B)
CSF-to-unbound serum concentration ratio at steady state.
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Analysis of the efflux clearance from the CSF after an
intracerebroventricular bolus administration.
The
CLCSF is given as a function of Q,
PSCSF, PSCSF,eff,
Dt and PSBBB,eff
(equation 23). By substituting the fitted parameter values to equation
23, CLCSF was calculated. As shown in figure 9, a fairly good agreement between
observed and fitted values was obtained for
CLCSF. Figure 9 also indicates that the diffusion through the brain tissue and the subsequent efflux across the BBB plays
an important role in the elimination of quinolones from the CSF after
intracerebroventricular administration.
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Fig. 9.
Contribution of the CSF bulk flow rate (striped
bars), active efflux clearance across the BCSFB (cross-hatched bars),
symmetrical permeability clearance across the BCSFB (open bars) and
diffusion through the brain tissues and the resultant efflux across the BBB (hatched bars) to apparent CSF efflux clearance after
intracerebroventricular bolus administration. Observed values of the
apparent CSF efflux clearance were taken from a previous report (Ooie
et al., 1996b) and represented as closed bars.
|
|
| |
Discussion |
The drug concentration in the brain ISF is a determinant for
in vivo CNS effects, although several processes needed to be considered for the precise analysis. Because there is free ligand exchange between CSF and ISF, the distributed model should be suitable
for the kinetic analysis of drug distribution in the brain tissue and
CSF (Collins and Dedrick, 1983; Suzuki et al., 1997). The
major assumptions of this model are that there is free ligand exchange
between CSF and ISF, and that the ligand molecules diffuse through the
brain parenchyma according to Fick's law. Both these assumptions have
been confirmed as being justified by previous reports in which the
ligand concentration in the brain parenchyma was determined as a
function of the distance from the ependymal surface in the
ventriculocisternal perfusion experiments. The kinetic analysis of the
experimental data revealed that the CNS profiles can be described by
the assumptions described previously (Patlak and Fenstermacher, 1975;
Blasberg et al., 1975; Fenstermacher and Davson, 1982). In
addition, Dykstra and his collaborators (1993) confirmed this
hypothesis by examining the brain concentration profiles after ligand
administration through a microdialysis probe implanted into the
cerebral cortex. Fenstermacher and Kaye (1988) summarized the
Dt values determined by the method
described previously and found a good relationship between
Dt and Dw.
Based on this relationship, we estimate the Dt
values for quinolone antibiotics.
The efflux clearance across the BBB (the sum of
PSBBB and PSBBB,eff) was
12- to 260-fold greater than PSBBB. Moreover,
after i.v. bolus administration, the concentration gradient was
observed from CSF to brain ISF; the brain acts as a sink (fig. 5).
These results provide kinetic evidence to support the hypothesis that a
significant efflux transport from the brain ISF to the capillary lumen
across the BBB is the predominant factor involved in the restricted
distribution of quinolones in brain ISF after systemic administration
(Ooie et al., 1997). Furthermore, the good correlation between PSBBB/(PSBBB + PSBBB,eff) and the
Kp,u,br at the steady state indicated that
the transport across the BBB is a determinant for the different
distribution of these quinolones in the brain tissue.
To evaluate the effect of drug diffusion through the brain tissue on
the apparent distribution in brain tissue and the CSF, simulation
studies were performed for the prediction of
Kp,u,br and
Kp,u,CSF at steady state assuming no drug
diffusion through brain tissue. As shown in figure 7, 16% to 52%
reduction was observed in Kp,u,br whereas
170% to 45% increase was demonstrated in
Kp,u,CSF. These results suggest that the
CSF acts as a pool to supply quinolone molecules to the brain ISF by
diffusion, whereas brain ISF acts as a sink to eliminate quinolone
molecules from the CSF at steady state following i.v. administration.
Regarding the efflux across the BCSFB, we have previously demonstrated
that the in vitro uptake clearance of -lactam
antibiotics by the isolated choroid plexus is similar to the in
vivo efflux clearance across the BCSFB determined by
intracerebroventricular administration (Ogawa et al., 1994).
We have also found that quinolone is transported by the isolated
choroid plexus via an organic anion transport system which
is also responsible for the efflux of -lactam antibiotics across the
BCSFB (Ooie et al., 1996a). Therefore, in the present study,
we have assumed that PSCSF,eff of quinolones is
the same as the uptake clearance determined by the isolated choroid
plexus reported previously (Ooie et al., 1996c). Assuming that the PSCSF,eff is zero, the participation of
active efflux transport across the BCSFB was studied by examining the
apparent distribution in the brain tissue and the CSF. As shown in
figure 8, active efflux transport across the BCSFB plays only a minor role in the distribution of quinolones both in brain tissue and the
CSF. These results are in good agreement with the results shown in
figure 9 that less than 20% of the apparent efflux clearance from the
CSF is caused by active efflux transport across the BCSFB. Unlike
-lactam antibiotics reported previously (Ogawa et al., 1994), these simulation results also agree well with our previous finding that probenecid has no significant inhibitory effect on the
apparent elimination of FLRX from the CSF after intracerebroventricular administration and that the elimination of FLRX was not saturable (Ooie
et al., 1996b).
As a barrier function of brain capillary endothelial cells, it has been
reported that P-glycoprotein acts as a primary active efflux transport
system to pump vincristine (Tsuji et al., 1992), cyclosporin A (Tatsuta et al., 1992) and vinblastine
(Schinkel et al., 1994) out of the brain into the
circulating blood. Moreover, several reports have provided kinetic
evidence that an efflux transport system for -lactam antibiotics
(Matsushita et al., 1991), azidodeoxythymidine (Dykstra
et al., 1993; Takasawa et al., 1997) and valproic
acid (Adkison et al., 1994) is located on the BBB. Although
this transport system has not been characterized yet, the transporter
may be different from P-glycoprotein (mdr1).
Regarding the CNS toxicity of quinolone antibiotics, drug
concentrations in the brain ISF would be a determinant for their duration and severity. The present study suggests that the affinity of
quinolones to a putative carrier-mediated efflux transport system may
be important in reducing brain ISF concentrations and consequent CNS
effect. Although a 10-fold difference was observed between SPFX and
OFLX for PSBBB,eff (table 3), comparison with the
physicochemical nature of quinolones revealed that there is no
significant relationship between the lipophilicity and
PSBBB,eff (fig. 4C). Because it has not been
possible to demonstrate the efflux transport of quinolones in the
primary cultured bovine brain capillary endothelial cells (Jaehde
et al., 1993), presumably because of the down-regulation of
the expression of the transporter, it would be important to use an
in vivo method to characterize the transport property across
the BBB. One of the rational ways to characterize the efflux transport
system at the BBB is to examine the elimination of ligand from the
brain after microinjection into the cerebral cortex (Leininger et
al., 1991; Banks et al., 1994; Kakee et al.,
1996).
In conclusion, it has been demonstrated that the significantly limited
distribution of quinolones in the brain tissue is caused by the
presence of an active efflux transport system located on the BBB.
Depending on the distance from the ependymal surface, there is a
significant concentration gradient from the CSF to the brain ISF after
systemic administration. One of the most important factors regulating
brain ISF concentrations of quinolones may be an affinity for the
putative efflux transport system on the BBB.
| |
GLOSSARY |
| Cp,u |
|
= |
|
unbound serum concentration |
| fu |
|
= |
|
serum unbound fraction |
| AUCu |
|
= |
|
area under the unbound serum concentration-time curve |
| PSBBB,app |
|
= |
|
apparent influx clearance across the BBB |
| PSCSF,app |
|
= |
|
apparent influx clearance across the BCSFB |
| PSBBB |
|
= |
|
symmetrical permeability clearance across the BBB |
| PSBBB,eff |
|
= |
|
efflux clearance across the BBB |
| PSCSF |
|
= |
|
symmetrical permeability clearance across the BCSFB |
| PSCSF,eff |
|
= |
|
efflux clearance across the BCSFB |
| Cbr |
|
= |
|
total brain concentration |
| CISF |
|
= |
|
ISF concentration |
| CCSF |
|
= |
|
CSF concentration |
| Kp,u,br |
|
= |
|
total brain-to-unbound serum concentration ratio
(Cbr/Cp,u) |
| Kp,u,CSF |
|
= |
|
CSF-to-unbound serum concentration ratio
(CCSF/Cp,u) |
| CLCSF |
|
= |
|
apparent efflux clearance from the CSF |
| Dt |
|
= |
|
diffusion coefficient in the brain tissue |
| Dw |
|
= |
|
diffusion coefficient in the agar |
| Mw |
|
= |
|
molecular weight |
| Vbr |
|
= |
|
distribution volume in brain tissue
(Cbr/CISF) |
| VCSF |
|
= |
|
volume of the CSF |
| Ar |
|
= |
|
surface area of the ependyma |
| Q |
|
= |
|
bulk flow rate of the CSF |
| L |
|
= |
|
thickness of the brain cortex |
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Abstract
Introduction
-aminobutyric acid;
CSF, cerebrospinal fluid;
ISF, interstitial
fluid;
BBB, blood-brain barrier;
BCSFB, blood-CSF barrier;
NFLX, norfloxacin;
OFLX, ofloxacin;
FLRX, fleroxacin;
PFLX, pefloxacin;
AM-1155, (±)-1-cyclopropyl-6-fluoro-1,4-dihydro-8-methoxy-7-(3-methyl-1-piperazinyl)-4-oxo-3-quinolinecarboxylic
acid ;
SPFX, sparfloxacin;
HPLC, high-performance liquid
chromatography.
-azido-3
