| Summary: | 博士 === 高雄醫學院 === 醫學研究所 === 87 === Abstract
Clozapine, an atypical antipsychotic agent with multiple receptor antagonist activities, was investigated in vivo and in vitro to discover its effects on intraocular pressure and blood flow, on the contractility of ciliary muscle and nonophthalmologic blood vessels, on cyclic AMP intermediation.and on calcium concentration in A7r5 smooth muscle cells. The adrenergic and muscarinic mechanisms involved in the above effects were also examined. In rabbits, clozapine (0.1, 0.25, and 0.5%) prolonged IOP recovery time and inhibited IOP response. Clozapine (0.1 and 0.25%) also produced a significant increase in ocular blood flow in this iris, ciliary, retina, and choroid at 30 to 180 min after drug administration. In isolated guinea pig thoracic aorta, clozapine relaxed phenylephrine (PE, 10μM)- and KCl (75mM)-induced contractions, the estimated IC50 values being 20.4±3.1 nM and 10.6±1.8 μM, respectively. Clozapine (0.1-100μM) inhibited phenylephrine (10μM)-induced influx of Ca2+, the estimated IC50 values being 0.4±0.1 nM, In isolated pig eye ciliary muscles, clozapine (1.0-100μM) inhibited carbachol (10μM)-induced contractions, the estimated IC50 value being 5.8±1.2μM. Clozapine (0.1-100μM) increased cyclic AMP, accumulation in pig''s ciliary bodies, including ciliary process, ciliary muscle, and trabecular meshwork. Pretreatment with carbachol (100μM) first decreased, then increased, clozapine-induced cyclic AMP accumulation. Studies of pretreatments with various muscarinic receptor antagonists at 100μM revealed that pirenzepine significantly enhanced clozapine (100μM)-induced cyclic AMP accumulation in trabecular meshwork, while 4-DAMP inhibited it in ciliary body, and methoctramine decreased it in ciliary process.
Clozapine elicited acute mild negative chronotropic effects and slow mild positive chronotropic effects in rat. Pretreatment with propranolol abolished the positive responses and retained the negative responses, pretreatment with PE abolished the negative responses and retained the positive responses. Pretreatment PE inhibited the hypotensive effect of clozapine.
Reserpinized and some nonreserpinized isolated spontaneously beating guinea pig right atrial preparations were used as the experimental model. Clozapine and acetycholine (ACh) decreased the spontaneously beating rate in a concentration-dependent manner at concentration up to 10μM. At lower concentration (5μM), clozapine increased the negative chronotropic effect of ACh, but at higher concentration (10μM), like atropine, restored the beating rate to the predrug control level. Pretreatment with atropine (50μM) and propranolol (10μM) increased the negative chronotropic effect of clozapine. Pretreatment with 3,4-DAMP (50μM), reduced the negative chronotropic effect of clozapine, but did not blocked it.
In coronary arteries, clozapine (0.1μM-100μM) inhibited contractions induced by KCl (35mM)，ouabain (10μM), and serotonin (10μM). In contrast, ACh (0.1-10μM) produced concentration-dependent constrictions. In Ca2+-free solution, the effect of clozapine was decreased while it was contracted by 35 mM KCl，but clozapine potentially inhibited the contractions induced by 4-AP, TEA, BaCl2 and ouabain. In Ca2+-free solution and endothelium denuded preparation, two phasic effects of clozapine was produced. Clozapine contracted the vessels in a concentration dependent manner at concentration up to 1μM, but at higher concentrations above 10μM , restored the contraction to the predrug control level. Pretreatment with atropine and 3,4-DAMP abolished the contraction but not relaxation. However, ACh ( 0.1-10μM) produced contraction in a concentration-dependent manner which was inhibited by clozapine. At lower concentrations, clozapine stimulated the receptors directly, and at higher concentration, it acted as an antimuscarinic agent. It showed significant difference before and after removal of endothlelium and extracellular Ca2+.
Pig coronary artery segments stored at 2-4℃ for 6 h in K+-free Krebs bicarbonate solution developed spontaneous rhythmic contractile response to [K+]o 25-40 mM, while cumulative additions of KCl 5 mM. Clozapine, both at lower concentration(<10nM) and at higher concentration (>1μM), block the rhythmic contraction, which may be attributed to fluctuation in intracellular calcium. Based on the contractilities in artery, it is suggested that at lower concentration (<10nM), clozapine showed moreα1 antagonist activity than muscarinic agonist activity, while at higher concentration (>1μM), it possesses more reversed muscarinic agoinst or muscarinic antagonist activity than muscarinic agoinst activity.
The ocular hypotensive effect of clozapine may be mostly attributed to its muscarinic agonist/antagonist activities and associated cyclic AMP increasing activities, which lead to ciliary muscle relaxation and a possibly associated increase in uveoscleral outflow. In cardiac studies, clozapine induced negative chronotropic effect. At lower concentration, clozapine increased the ACh-induced negative chronotropic effect, but at higher concentration, it restored the beating rate to near predrug control level. However, in the coronary studies, the contraction induced by lower concentration of clozapine could be changed into relaxation at higher concentrations. Pretreatment with atropine and 3,4-DAMP abolished the contraction but not relaxation. The biphasic effects proved that clozapine possessed muscarinic agonist and antagonist effects on ocular and cardiovascular muscarinic ACh receptors.
Clozapine’s ability to increase blood flow and relax vessels may be attributed to its ability to blockα1-adrenoceptor. Clozapine, with blood flow increasing activities, produced fewer side effects such as ischemic retinophathy, when used to decrease IOP. IOP are considered useful for glaucoma treatment. Based on the results above, clozapine may potentially be important in the development of new antiglaucoma agents.
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