Download Therapeutic and systemic side effects of ocular beta

Therapeutic and Systemic Side Effects of Ocular
/?-Adrenergic Antagonists in Anesthetized Dogs
Alan L. Svec and Arthur M. Strosberg
Beta adrenergic antagonists, while useful in the treatment of glaucoma, can be absorbed transocularly
causing a variety of systemic side effects. An adequate animal model which simultaneously indicates
the intraocular pressure lowering effects as well as the systemic effects of ocularly applied 0-adrenergic
antagonists would be highly useful. The pentobarbital anesthetized dog, instrumented to record blood
pressure (BP) and heart rate (HR) and in which intraocular pressure (IOP) was measured with a pneumatonometer, was investigated. Isoproterenol iv dose-response curves were run and IOP measured before
and hourly for 3 hr following ocular application of 50 n\ 0.5% timolol, 0.5% RS-52367 (a Syntex
/9-adrenergic antagonist), or dH2O control. Timolol and RS-52367 produced similar decreases in IOP.
However, timolol decreased basal BP and HR, markedly inhibited isoproterenol-induced BP and HR
responses, and also decreased contralateral IOP. Systemic effects of RS-52367 were far more mild.
Since both the IOP lowering and systemic /3-adrenergic antagonist properties of timolol were elicited,
the anesthetized dog appears to be an excellent animal model for examining the effects of /8-adrenergic
antagonists applied topically to the eye. Invest Ophthalmol Vis Sci 27:401-405, 1986
a j8,-selective Syntex /3-adrenergic antagonist5) on ipsilateral and contralateral IOP, baseline blood pressure
and heart rate, and on isoproterenol-induced blood
pressure and heart rate changes (as an indicator of /3antagonism) were determined.
/3-adrenergic antagonists are useful in the treatment
of open angle glaucoma by lowering intraocular pressure (IOP)1 via reduction of the rate of formation of
aqueous humor. 2 Clinical studies reveal that timolol
maleate, a /3-adrenergic antagonist shown to be effective
in lowering IOP in glaucoma patients, is absorbed systemically following ocular application and causes a variety of side effects.3 Of particular significance are the
cardiovascular effects, which may intensify failure in
congestive heart failure patients, and the pulmonary
tract jS-adrenergic antagonist effects, which may lead
to bronchospasm in asthmatics or patients with pulmonary obstructive disease.4
One of the major difficulties in determining the IOP
lowering efficacy and potential for systemic side effects
of /3-adrenergic antagonists is the lack of a suitable experimental animal model. The present study was performed to determine the utility of the pentobarbital
anesthetized dog as an experimental animal model for
evaluating both the IOP lowering ability and cardiovascular side effects of/3-adrenergic antagonists applied
topically to the eye. The effects of ocularly administered
timolol and RS-52367 (l-t-butylamino-3[5'-5 methyl
hexyl aminocarbonyl)-2-thiazoloxy]propan-2-ol HC1,
Materials and Methods
Experimental Preparation
Mongrel dogs of either sex (12.6 kg ± 0.8 kg) were
anesthetized with pentobarbital Na iv (30 mg/kg) and
supplemented by continuous iv infusion (5 mg/kg/hr).
A femoral vein was cannulated for injection and a
femoral artery for blood pressure measurement. Heart
rate was recorded by a cardiotachograph triggered by
the R wave of a limb lead II ECG. Responses were
recorded on a Beckman type R polygraph (Sensor
Medics Corp., Anaheim, CA). Intraocular pressure
measurements were made with a Digilab model 30-D
pneumatonometer (Digilab; Cambridge, MA). Respiration was maintained constant with a respirator (Harvard Apparatus; So. Natick, MA).
Experimental Protocol
Isoproterenol was administered iv in half-log doses
from 0.0158 Mg/kg to 0.5 Mg/kg at 10-min intervals
(pre-drug control). A 30-min rest period followed. Single 50 n\ drops of 0.5% Timoptic (Merck, Sharp, &
Dohme; West Point, PA), 0.5% RS-52367, or dH 2 O
were then instilled into the inferior conjunctival sac of
one eye with a micropipette. Isoproterenol dose-response curves were again run hourly for 3 hr. Ipsilateral
From the Institute of Pharmacology and Metabolism, Syntex Research, Palo Alto, CA 94303.
Presented in part as a poster at the 9th International Congress of
Pharmacology, London, August 1984.
Submitted for publication: May 15, 1985.
Reprint requests: Arthur M. Strosberg, Syntex Research, 3401
Hillview Avenue, Palo Alto, CA 94303.
401
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402
INVESTIGATIVE OPHTHALMOLOGY & VISUAL SCIENCE / March 1986
"*"•-.«,
0
30
60
90
TIME
I
120
dMIO
150
I
180
(min)
Fig. 1. IOP* response in ipsilateral eyes of anesthetized dogs treated
with single 50-^1 drop ocular applications of 0.5% timolol
(•
• ) , or 0.5% RS-52367 ( •
A ) , as compared to dH 2 O
• ) . Each point is the mean ± SEM, n = 5: *P < 0.005,
control ( •
"I*/* < 0.01. *IOP is expressed in mm Hg equivalents.
IOP was measured immediately before administration
of drug, and again hourly, prior to each isoproterenol
dose response curve. Blood pressure and heart rate were
monitored throughout the experiment.
Studies employing the same protocol were also performed in similarly instrumented dogs to determine
the effect of each compound on contralateral IOP following ocular administration.
In additional studies utilizing the same protocol except for the route of administration, the effects of 50
ixl intravenous injections of 0.5% timolol and 0.5% RS52367 were examined in similarly instrumented preparations.
Data were compared for statistical significance using
Student's t-test.6
O
30
60
Vol. 27
90
TIME (min)
120
150
130
Fig. 2. IOP* response in ipsilateral (A
A), and contralateral
eyes ( •
• ) of anesthetized dogs treated with single 50-jtl drop
ocular applications of 0.5% timolol. n = 3, as compared to dH 2 O
control, n = 5 ( •
• ) . Each point is the mean ± SEM; *P < 0.025,
iP < 0.1. *IOP is expressed in mm Hg equivalents.
Fig. 3. Change in mean arterial blood pressure in anesthetized
dogs treated with single 50-MI drop ocular applications of 0.5% timolol
(•
• ) , or 0.5% RS-52367 (A
A ) , as compared to dH 2 O
control ( •
• ) . Each point is the mean, n = 6: *P < 0.05. Standard error bars are omitted for clarity.
Drug Preparation
RS-52367 was dissolved in dH 2 O to a concentration
of 0.5% Timoptic solution 0.5% (timolol maleate MSD,
Lot. No. 138OJ) was obtained commercially. Isoproterenol (Aldrich Chemicals; Milwaukee, WI) was dissolved in saline containing ascorbic acid as a preservative. All solutions were computed as free base concentration.
These studies adhere to the ARVO Resolution on
the Use of Animals in Research.
Results
Intraocular Pressure
The effects of ocular 0.5% timolol, 0.5% RS-52367
and dH 2 O on IOP in treated eyes are shown in Figure
1. Both compounds produced statistically significant
(P < 0.005), similar reductions in IOP as compared to
dH 2 O control for 3 hr following a single 50-^1 ocular
application. At 3 hr, however, RS-52367 affected a
slightly greater reduction in IOP than timolol. Mild
irritation of the conjunctiva and sclera of treated eyes
was noted in some dogs receiving RS-52367.
The effects of 0.5% timolol on IOP in contralateral
eyes as compared to treated eyes and dH 2 O control are
shown in Figure 2. Timolol produced statistically significant (P < 0.025) reductions in ipsilateral IOP as
compared to dH 2 O control for 3 hr following a single
50-^1 ocular application. Contralateral IOP exhibited
a tendency towards reduction (P < 0.10) beginning 1
hr after dosing. The difference between ipsilateral and
contralateral IOP was not statistically significant (P
< 0.05) at 2 and 3 hr.
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No. 3
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OCULAR 18-ADRENERGIC ANTAGONISTS IN ANESTHETIZED DOGS / Svec and Srrosberg
Ocular application of 50 /*1 0.5% RS-52367 did not
produce a reduction in contralateral IOP.
Effects on Blood Pressure and Heart Rate
The effects of ocular 0.5% timolol, 0.5% RS-52367,
and dH2O control on baseline blood pressure and heart
rate are shown in Figures 3 and 4. Timolol significantly
lowered blood pressure and heart rate (P < 0.05) within
30 min after ocular application. Blood pressure and
heart rate returned to near pre-drug levels at 3 hr. Intravenous timolol produced similar decreases in blood
pressure and heart rate. However, these effects persisted
at 3 hr.
Blood pressure and heart rate in dogs treated with
ocular RS-52367 were not significantly different from
dH2O controls (P > 0.05) over the 3-hr time period
(Figs. 3,4). Intravenous RS-52367 produced no change
in blood pressure. A decrease in heart rate was noted
in two animals receiving intravenous RS-52367. This
decrease began 1 hr after dosing and did not correlate
with any associated isoproterenol blockade.
Inhibition of Isoproterenol Responses
Ocular timolol markedly inhibited iv isoproterenolinduced blood pressure and heart rate responses at 1,
2 and 3 hr after administration (Table 1). The same
dose of timolol given intravenously produced a similar
inhibition (Table 2). These data indicate that upon
ocular application timolol is readily absorbed systemically in the dog and produces significant systemic /3adrenergic antagonism.
In contrast, ocular RS-52367 produced only slight
inhibition of isoproterenol challenges (Table 1). Studies
with intravenous RS-52367 at 50 n\0.5% solution (approx. 20 Mg/kg) showed the compound possessed little
60
90
120
ISO
180
TIME (min)
Fig. 4. Change in heart rate in anesthetized dogs treated with single
50-/zl drop ocular applications of 0.5% timolol ( •
• ) or 0.5%
RS-52367 ( •
A), as compared to dH 2 O control ( •
• ) . Each
point is the mean, n = 6: *P < 0.05. Standard error bars are omitted
for clarity.
isoproterenol blocking activity at this dose (Table 2).
However, higher doses (>316 Mg/kg) produce a complete antagonism of isoproterenol induced cardiac responses with little effect on blood pressure responses.5
Discussion
The systemic absorption and resultant side effects
of ocularly applied agents has long been a problem in
drug therapy.7 Topically applied /3-adrenergic antagonists, specifically timolol, are quite effective in treating
open angle glaucoma by lowering IOP. However, timolol is contraindicated for this use in patients with
bronchial asthma, obstructive pulmonary disease, second and third degree heart block, sinus bradycardia
and cardiac failure.8 This is because the same cardiac
and pulmonary effects seen with systemically admin-
Table 1. Effects of ocular timolol* and RS-52367* on isoproterenol-induced blood pressure
and heart rate responses in anesthetized dogs
Percent inhibition of pre-drug isoproterenol responsef
Hour I
Isoproterenol
(tig/kg, iv)
Hour 2
Hour 3
Response
Timolol
RS-52367
dH2O
Control
0.0158
mean BP
heart rate
97.0 ± 1.6*
96.7 ± 3.3*
0.0 ± 0.0
15.8 ±7.8
1.5 ± 1.5
0.0 ± 0.0
94.2 ± 1.5*
89.7 + 6.0*
11.5 + 5.9
13.2 ±7.4
0.7 ± 0.7
0.0 ± 0.0
88.0 ± 4.2*
83.6 ± 5.3*
14.7 ± 8.8
28.2 ± 11.8
3.7 ± 2.9
6.5 ± 5.9
0.05
mean BP
heart rate
93.5 ± 2.2*
87.7 ± 4.4*
1.1 ± 1.1
17.0 ±5.7
3.0 ± 3.0
3.8 ± 2 . 1
86.5 ± 2.7*
81.5 ± 6.2*
18.3 ±2.7
21.7 ±7.3
3.0 ± 3.0
4.8 ± 2.8
70.2 + 5.8*
72.2 ± 6.1*
23.8 ± 11.9
38.3 ± 8.2*
5.5 ± 5.5
8.3 ± 4.7
0.158
mean BP
heart rate
76.3 ± 5.8*
75.8 ± 7.0*
0.3 ± 0.3
10.2 ± 3.0
2.2 + 2.2
1.8 ±2.1
66.0 ± 6.5*
64.0 ± 8.3*
13.8+9.0
18.2 ±8.3
1.8 ± 1.8
4.7 ± 2.3
55.3 ± 7.0*
57.0 ± 8.1f
17.8 ± 10.8
20.8 ± 7.4
3.2 ± 3.2
10.3 ± 3.5
0.5
mean BP
heart rate
60.8 ± 5.7*
57.6 ± 14.0*
0.4 ± 0.4
10.2 ± 3.5
2.0 ± 1.6
1.0 ± 1.0
50.2 ± 6.5*
43.8 ± 12.4
9.0 ± 7.0
11.5 + 3.6
1.0 ± 1.0
2.8 ± 1.8
39.0 ± 5.3*
37.0 ± 10.8
9.2 ± 6.9
13.3 ± 3.4
0.3 ± 0.3
8.2 ± 3.7
* 50 nl, 0.5% solution, ocular administration, as compared to 50 ^1 dH2O.
Timolol
RS-52367
dH2O
Control
Timolol
RS-52367
dH2O
Control
* / > < 0.001
t mean ± SEM, n = 6
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404
INVESTIGATIVE OPHTHALMOLOGY 6 VISUAL SCIENCE / March 1986
Vol. 27
Table 2. Effects of intravenous timolol and RS-52367 on isoproterenol-induced blood pressure
and heart rate responses in anesthetized dogs
Percent Inhibition ofpre-drug isoproterenol response
Hour 1
Isoproterenol Response
Hour 2
Hour 3
Timolol*
RS-52367*
Controlf
Timolol*
RS-52367*
Controlf
Timolol*
RS-52367*
Controlf
0.0158
mean BP
heart rate
100.0 ± 0.0$
98.0 ± 2.0$
9.3 ± 9.3
22.0 ± 11.1
1.5 ± 1.5
0.0 ± 0.0
100.0 ± 0.0$
98.0 ± 2.0$
10.3 ± 10.3
3.3 ± 3.3
0.7 ± 0.7
0.0 ± 0.0
98.7 ± 1.3$
94.3 ± 5.2$
11.0± 11.0
17.0 ± 17.0
3.7 ± 2.9
6.5 ± 5.9
0.05
mean BP
heart rate
98.0 ± 1.2$
93.0 ± 3.0$
3.6 ± 3.6
15.3 ± 7.8
3.0 ± 3.0
3.8 ±2.1
92.7 ± 1.8$
90.3 ± 2.7$
7.3 ± 7.3
11.3± 9.4
3.0 ± 3.0
4.8 ± 2.8
90.3 ± 2.3$
83.3 ± 4.7$
9.0 ± 6.7
19.0 ± 16.0
5.5 ± 5.5
8.3 ± 4.7
0.158
mean BP
heart rate
93.0 ± 2.3$
83.0 ± 5.6$
0.0 ± 0.0
8.7 ± 8.7
2.2 ± 2.2
1.8 ±2.1
84.8 ± 5.8$
75.0 ± 8.2£
0.0 ± 0.0
8.0 ± 8.0
1.8 ± 1.8
4.7 ± 2.3
77.0 ± 5.0$
67.3 ± 1.6$
0.0 ± 0.0
12.0 ± 11.0
3.2 ± 3.2
10.3 ±3.5
0.5
mean BP
heart rate
80.0 ± 4.2$
64.3 ± 9.5$
1.0 ± 1.0
8.3 ± 4.6
2.0 ± 1.6
1.0 ± 1.0
68.0 ± 5.0£
51.6 ± 16.4£
2.3 ± 2.3
9.0 ± 4.5
1.0± 1.0
2.8 ± 1.8
57.6 ± 8.1f
46.0 ± 19.7
4.0 ± 4.0
10.3 ± 5.2
0.3 ± 0.3
8.2 ± 3.7
* 50 pi, 0.5% solution, intravenous administration; mean ± SEM, n = 3.
t 50 fi\ dH2O ocular administration; mean ± SEM, n = 6.
istered /?-adrenergic antagonists, viz, cardiac depression
and antagonism of fo receptor mediated bronchodilation, can be produced with ocularly applied timolol,3
indicating the compound is absorbed transocularly. In
rare instances, death by cardiac failure has occurred
following ocular timolol administration. 4
As with many therapeutic agents, a relatively uncommon side effect may not be clearly identified in
clinical testing. A statistically significant number of adverse effects show up only when the agent is given to
a large enough population of susceptible individuals.
With timolol, the cardiovascular and pulmonary side
effects were found to be inconspicuous except in patients with pre-existing cardiovascular or pulmonary
disease states.9 As the popularity of using ocularly applied /3-adrenergic antagonists to treat open angle glaucoma grows, so does the potential for occurrence of
deleterious effects produced by newer agents. These
agents will, by default, have had less documentation
concerning exposure to a susceptible population. An
animal model which simultaneously depicts the IOP
lowering effects of /3-adrenergic antagonists as well as
their potential for cardiovascular side effects would be
highly useful in the early identification of compounds
which lower IOP but produce minimal systemic side
effects.
Several animal models including normotensive10 and
water loaded albino rabbits,11 cats,12 and cynomolgus
monkeys13 have been employed to assay the effects of
ocularly applied /^-antagonists but have met with mixed
success. For example, Liu et al, demonstrated a lowering of IOP with timolol applied to the eyes of waterloaded rabbits. However, this did not correlate with
decreases in heart rate and blood pressure induced by
timolol in these animals.14
$ P < 0.0025.
£ P < 0.05.
The present study has demonstrated the utility of
the pentobarbital anesthetized dog as a model for the
study of ocularly administered /?-adrenergic antagonists. In this model, ocular timolol produced decreases
in IOP similar to those seen in man. 1 The typical cardiovascular side effects seen in man with timolol 4 were
also elicited in the dog. The marked antagonism of
isoproterenol induced blood pressure and heart rate
responses and a tendency toward some lowering of
contralateral IOP following ocular timolol administration confirm the systemic absorption of this agent.
The Syntex compound RS-52367 appears useful in
lowering IOP. While the compound is less potent than
timolol as a /3-adrenergic antagonist by intravenous
administration, it produces an equivalent lowering of
IOP when identical doses are applied ocularly. This
may be due to a local high concentration of RS-52367
relative to timolol if RS-52367 is systemically absorbed
to a lesser degree than timolol or if RS-52367 is absorbed into the anterior chamber of the eye to a greater
degree than timolol. If RS-52367 can be formulated
in a nonirritating preparation, the compound may be
safe and effective in the treatment of open angle glaucoma.
The pentobarbital anesthetized dog has been used
for many years in cardiovascular pharmacology research and remains an accurate model for studying
many of the physiological and pharmacological characteristics of man. The adaptation of this model to
ocular research will hopefully speed the development
of safe and effective new drugs to treat open angle glaucoma.
Key words: /3-adrenergic antagonists, timolol, glaucoma, cardiovascular, intraocular pressure
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No. 3
OCULAR 0-ADRENERGIC ANTAGONISTS IN ANESTHETIZED DOGS / Svec and Srrosberg
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