Biomedical Engineering Reference
In-Depth Information
(a)
Spike rate
Burst rate
SO121A, FC, 46div, 45 units
1200
100
W
REF
Muscimol titration in presence of 20
µ
M gabazine
90
80
70
60
50
40
30
20
10
0
1000
10
20
50
70
120
160
30
800
600
400
200
0
0
30
60
90
120
150
180
210
240
270
300
330
Time (min)
(b)
(c)
2.8
100
Slope =
1.29 ± 0.23
2.4
80
10 µM, n=2
20 µM, n=4
40 µM, n=2
2.0
60
1.6
40
pA2
= 6.6
1.2
Gabazine (
µ
M)
0.8
20
0
10
20
40
0.4
0
0.0
1E-4
1E-3
0.01
0.1
1
10
100
1000
-7.0
-6.5
-6.0
-5.5
-5.0
-4.5
-4.0
Muscimol (
µ
M)
Log gabazine (M)
FIGURE 6.10
Quantification of receptor interactions for the GABA antagonist gabazine. (a) Muscimol titration in the presence
of 20 µM gabazine. Each data point represents average spike rate (left ordinate) and burst rate (right ordinate)
for all discriminated units (45) in bins of 1 min. Note level response activity plateaus from which percent decrease
from reference activity can be determined. (b) Concentration-response curves of the pooled data in the presence
of 0, 10, 20, 40 µM gabazine. Vertical bars represent SD of percent spike activity decrease. The mean IC 50 for mus-
cimol in the presence of gabazine are 20.1, 37.2, and 120.4 µM. (c) Schild plot of log (dose ratio-1) vs. log gabazine
concentration. The X -intercept represents a p A 2 value (-log of K B ) of 6.6. The dissociation constant K B is 0.25 µM
(redrawn from Oli-Rijal, 2005).
mechanism for muscimol, its concentration in medium remains constant and it is well
suited for quantitative pharmacology of the GABA A receptor. Gabazine, [2-(3'-carboxy-2'-
propyl)-3-amino-6-p-methoxyphenylpyridazinium bromide], is an aryl-aminopyridazine
derivative of gamma-aminobutyric acid (GABA) and represents a potent, selective, com-
petitive, and reversible GABA A antagonist [21, 22]. Muscimol titration in the presence of
increasing concentrations of gabazine shifts the concentration-response curves (CTCs) to
the right (higher concentrations of muscimol), but with identical efficacies and with no
measurable change in slopes (Figure 10a). A Schild plot [23]) with linear regression pre-
dicts an X -intercept of 0.23 µM. This intercept is the dissociation constant K B for the antag-
onist gabazine. The theoretical slope of 1 identifies a substance as a competitive
antagonist. The experimental slope of 1.29
0.23 is not different from unity and identifies
gabazine as a competitive antagonist of muscimol. The K B and the p A 2 value (-log K B ) of
6.63 agree well with published data [24, 25] and validate primary cultures as useful, his-
totypic biosensor candidates.
 
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