This class of drugs does not play a role in ophthalmology.

Cholinesterase inhibitors increase cholinergic activity in both muscarinic and nicotinic structures. With two exceptions, these drugs are used by the ophthalmologist for their muscarinic effects. One exception is their use in the diagnosis of myasthenia gravis.

Intravenous edrophonium (Tensilon) and, to a lesser extent, intramuscular neostigmine²²¹ have been the cholinesterase inhibitors used most frequently.

Edrophonium (Fig. 1) is a short-acting cholinesterase inhibitor. It carries a positive charge, enabling it to bind with the negatively charged “anionic” site of acetylcholinesterase. The bulky cyclic structure of edrophonium physically blocks the approach of acetylcholine to the “esteradic” site. In addition, electrostatic hydrogen bonding of the hydroxyl group of edrophonium causes inactivation of the esteradic site. Acetylcholine hydrolysis is inhibited because it must compete with edrophonium for both the “anionic” and “esteradic” sites. Edrophonium’s binding to both enzyme sites is weak and short lived.

The side effects of intravenous edrophonium are the result of its action on muscarinic structures (bowel, bladder, and heart). They include abdominal cramping, urinary incontinence, bradycardia, and cardiac arrest.^222,223 However, the diagnostic value of edrophonium in myasthenia gravis resides in its nicotinic action: the striated musculature contracts more vigorously. Therefore, the side effects of edrophonium can be prevented without interfering with the drug’s diagnostic efficacy by simultaneously administering a muscarinic blocking agent, such as atropine. This could be done as follows: Two syringes are used. One contains saline; the other contains atropine, 0.4 mg, and edrophonium, 10 mg. Neither the patient nor the physician evaluating the response knows which of the two solutions is being injected. The nurse gives one syringe to the physician for injection. Three tenths of the volume is injected rapidly, and the response is evaluated during the next minute. If this is well tolerated, the remainder is injected and the response is evaluated. The contents of the second syringe are then injected in the same manner. Only after the patient and physician have committed themselves to an evaluation of the relative efficacy of the two syringes does the nurse identify which contained the edrophonium.

Several different parameters have been suggested for evaluating the edrophonium response. The objective improvements in ocular and lid motility have been of primary interest to the ophthalmologist. Glaser and co-workers^224,225 used tonography to document the increased extraocular muscle tonus. They reported a mean 2-mmHg increase in intraocular pressure within 1 minute of injecting 10 mg edrophonium in 15 myasthenic patients. All 15 had unilateral, but not always bilateral, responses. The maximum elevation was 5 mmHg. The response could occur in less than 10 seconds, but the mean response interval was approximately 20 seconds²²⁶ to 35 seconds.²²⁷ Although there were no false-positive results in the 10 control patients in Glaser and co-workers’ study, false-positive results can occur.^227,228 Wray and Pavan-Langston²²⁸ found that the intraocular pressure increase was greater in myasthenic patients without ophthalmoplegia than in those with ophthalmoplegia.

Kornbleuth and colleagues²²⁹ used electromyographic recordings of the extraocular muscles when testing with edrophonium. In their small series, only patients with myasthenia gravis showed an increase in electrical activity. Campbell and associates²²⁷ used electronystagmography to study the effect of edrophonium on the fatigue induced by optokinetic nystagmus. A beneficial response occurred in 50% of 17 myasthenia gravis patients given edrophonium injections. There were no false-positive reactions among 18 normal subjects. However, these authors found the test to be of only limited value because of the marked variations in amplitude and rate of optokinetic nystagmus among subjects.

Edrophonium increases the peak saccadic velocity in myasthenic patients but decreases it in normal controls.²³⁰ The use of topical cholinesterase inhibitor drops in the treatment of ocular myasthenia gravis has been suggested,²³¹ but their value is not well documented.

Cholinesterase inhibitors are also used for their nicotinic effects in the treatment of pediculosis of the lashes. The cholinesterase inhibitors are pesticides and kill by producing respiratory arrest. Cholinesterase inhibitors used primarily for their intraocular effects are relatively ineffective pesticides. Physostigmine 0.1% in peanut oil and 0.25% ointment kill the adult form of the crab louse (Phithirus pubis), but the nits are more resistant and must be removed with forceps.^232,233 Malathion is more effective and relatively safe because it is rapidly inactivated by human serum carboxylesterases. Within 5 minutes of exposure, 100% of lice are killed, and after 10 minutes of exposure 96% of eggs will not hatch. Two problems encountered when using malathion around the eyes are that commercial preparations contain a high (e.g., 78%) alcohol content and the drug gives off a foul-smelling sulfhydryl degradation product.

Demodex folliculorum is another pest infesting the lids. It is quite resistant to ophthalmic preparations. It can survive 3 or more days in 0.125% echothiophate, 0.25% demecarium, or 0.1% diisopropyl fluorophosphate.²³⁴

Two classes of nicotinic blocking agents exist: nondepolarizing antagonists (e.g., curare) and depolarizing antagonists (e.g., succinyldicholine).

NONDEPOLARIZING ANTAGONISTS. Neuromuscular Blockade. Duke-Elder and Duke-Elder²³⁵ showed that acetylcholine and nicotine caused contraction of dog extraocular muscles and that curare, but not atropine, could block these responses. Muscarine was subsequently shown to have a very weak contractile effect that could be blocked by atropine.²³⁶ Until the work of Katz and Eakins,²³⁷ it was believed that the ocular muscles were more sensitive to the blocking effects of nondepolarizing agents, such as curare and gallamine, than were the peripheral skeletal muscles.²³⁸ Katz and Eakins found that the reverse seemed true (e.g., a systemic dose of curare that paralyzed the cat anterior tibial muscle produced only a 50% block of the twitch response of the superior rectus). Their work was supported by Sanghvi and Smith.²³⁶ Systemic administration of nondepolarizing relaxants, such as alcuronium,²³⁹ atracurium,^240,241 fazadinium,²⁴² metocurine, and pancuronium,²⁴³ is associated with a lowering of intraocular pressure. Some of these have a rapid onset and short duration of action, which makes them useful as induction agents for intubation. These drugs also differ in their cardiovascular effects. For example, alcuronium and curare lower arterial blood pressure, whereas pancuronium increases blood pressure and heart rate.²⁴⁴

An effective neuromuscular block with atracurium or vecuronium lasts approximately 30 minutes, making these agents intermediate in their duration of action. Atracurium, unlike the shorter-acting depolarizing antagonist succinyldicholine, undergoes rapid ester hydrolysis at physiologic pH independent of serum butyrylcholinesterase activity. Large doses of atracurium have been suggested as a substitute for succinyldicholine induction because the former does not produce extraocular muscle contraction.^245,246 Atracurium 0.5 mg/kg did not produce a significant rise in intraocular pressure in subjects under steady-state nitrous oxide general anesthesia, but succinyldicholine 1 mg/kg did, from a mean baseline of 5.6 mmHg to 13.2 mmHg. Vecuronium seems to act in a manner similar to that of atracurium.^247,248

Mivacurium, at a dose of 0.15 mg/kg, has a slower onset of action (approximately 2.5 minutes) than succinyldicholine. At this dose, mivacurium’s striated muscle block lasts approximately 16 minutes. Mivacurium, unlike atracurium and vecuronium, is inactivated primarily by butyrylcholinesterase.

Traction on the extraocular muscles during general anesthesia produces slowing of the heart. As stated earlier, pancuronium produces a tachycardia, which is protective. Strabismic children under general anesthesia who received pancuronium had heart rates that dropped from 120 ± 17 beats per minute to 94 ± 25 beats per minute.²⁴⁹ Patients receiving atracurium had pulse rates that fell from 86 ± 20 beats per minute to 34 ± 22 beats per minute, and 40% developed an additional arrhythmia besides the bradycardia. From a cardiac viewpoint, pancuronium has advantages in strabismus surgery.

Ganglionic Blockade. Transmission in sympathetic ganglia (e.g., superior cervical ganglion) and parasympathetic ganglia (e.g., ciliary ganglion) is primarily nicotinic. Several investigators have looked at ocular effects that might be related to partial blockade at this level. Gallamine²⁵⁰ and pancuronium²⁵¹ administration have been associated with a lowering of intraocular pressure. This may have been due, in part, to a reduction in systemic blood pressure. Intubation for general anesthesia produced tracheal irritation and a reflex increase in blood pressure. When throats were not anesthetized with topical anesthesia, the intraocular pressures 5 minutes after gallamine injection remained at baseline; when throats were anesthetized, the intraocular pressures were below baseline. The reductions in intraocular pressures produced by pancuronium were significant but were not dose related with the use of 0.01 to 0.08 mg/kg. Trimetaphan, a short-acting ganglionic blocking agent, administered intravenously produced reductions in intraocular pressure from preoperative values of 13 to 17 mmHg to intraoperative values of 3 to 4 mmHg as the systolic blood pressure fell to 60 mmHg.²⁵²

Hexamethonium and pentamethonium, 2 mg/kg, given intramuscularly to conscious subjects caused a mean maximum fall in intraocular pressure of 9 mmHg.²⁵³ The decline was present within 30 minutes of injection and persisted for 3 hours. This effect was attributed to a blockade of sympathetic ganglia and a resultant lowering of systemic blood pressure. Barnett²⁵³ treated hypertensive patients for 9 to 11 weeks with these drugs. He reported an improvement in the hypertensive retinopathy. However, these drugs produced an orthostatic hypotension that was disabling, which limited their use.

Retrobulbar tetraethylammonium injection in human subjects has produced mydriasis.²⁵⁴ The pupil remained dilated longer than 6 hours and did not constrict after topical physostigmine. Ciliary ganglion blockade also produced cycloplegia.^255,256 Decreased tearing has been attributed to blockade of the sphenopalatine ganglion after systemic administration of tetraethylammonium.²⁵⁵

Based on the assumption that sympathetic superior cervical and stellate ganglia fibers innervate and constrict the retinal arterioles, ganglionic blocking agents have been used in the treatment of central retinal artery occlusions, in the hope that the vessels would dilate and the embolus pass to a more distal arterial branch. However, there is little evidence that they are beneficial, and it can be argued that they may further reduce the perfusion blood pressure to the eye.

Botulinum Toxin. Botulinum toxins are produced by the anaerobic bacteria Clostridia botulinum. Each of eight different strains produces an immunologically distinct form of the toxin: A, B, C₁, C₂, D, E, F, and G. Four, A, B, D, and E, have been shown to block cholinergic function. They bind to unmyelinated cholinergic nerve terminals and prevent the release of quantal acetylcholine.^257,258 Death occurs by paralysis of the respiratory muscles; doses as low as 1 ng per kilogram body weight can be lethal in humans.²⁵⁹

Botulinum toxins are zinc-dependent proteases that attack proteins linking the acetylcholine-containing vesicles to the synaptic axon membrane. Calcium is no longer able to trigger a functional release of the transmitter.²⁸ The structure of botulinum A toxin has been fully sequenced.²⁶⁰ Type A botulinum toxin attacks a soluble 25-kd cytoplasmic attachment protein, SNAP-25,^29,261,262 abolishing quantal (i.e., functional) acetylcholine exocytosis; some nonquantal acetylcholine release does persist. The toxin does not block the neuron’s action potential, the synthesis or storage of acetylcholine, the vesicle number, or calcium entry.²⁶³ The initial binding site of botulinum A toxin is a sialic acid-containing site on the external neuron surface; this attachment can be antagonized by lectins.²⁶⁴ Once inside the neuron, botulinum toxin can no longer be neutralized by antitoxins.

Type A botulinum toxin is more effective at nicotinic neuromuscular junctions than at nicotinic ganglion synapses or parasympathetic neuromuscular junctions. It has been used to treat essential blepharospasm, blepharospasm associated with facial dyskinesias, hemifacial spasm, nonparalytic and thyroid ophthalmopathy-induced strabismus, paralytic strabismus, nystagmus, corneal exposure, and entropion.^{265,266,267,268,269,270,271,272,273,274,275,276,277,278}

The toxin is injected into the muscle to be rendered paretic or subcutaneously in the adjacent area. The effect is dose dependent. Multiple small doses (e.g., 0.001 to 0.003 μg; 2.5 to 7.5 U) or fewer but larger doses may be given. The onset of effect is within 48 hours, and the maximum effect may take 5 to 6 days to develop. The results are usually transient and average 2 to 3 months in duration.

Histologic examination of orbicularis muscle fibers after prolonged treatment of blepharospasm failed to show morphologic changes. However, the motor neurons had many unmyelinated axon sprouts, most of which ended blindly; those ending on muscle fibers were able to elicit new and multiple motor end plates at previously nonsynaptic sites.^279,280

In general, botulinum A treatment is of little value in strabismus in which the eye muscle movements are restricted, and it is of less value than traditional surgery in comitant deviations.²⁸¹ Retrobulbar injections, used to treat nystagmus and oscillopsia, may be of temporary value depending, in part, on the amount and duration of ptosis produced.^{282,283,284,285} Complications occur from local spread of toxin to adjacent muscle fibers and from an inability to accurately predict the magnitude of the effect. Ptosis is one of the most common complications caused by local spread of toxin.²⁸⁶ Ptosis, partial or complete, can at times be of therapeutic value (e.g., before recovery from Bell’s palsy).^287,288,289 Deliberate ptosis can be achieved by injecting botulinum A toxin along the superior orbital roof, approximately 25 mm behind the superior orbital rim.

Clinical signs of systemic toxicity have not been reported, but there is evidence of subtle systemic effects (e.g., abnormal electromyographic recordings in arm muscle fibers of patients receiving 0.005 μg of toxin).²⁹⁰ Neurologists treat dystonias with relatively large doses of botulinum A toxin; these may produce only subtle distant electromyographic changes but are likely to elicit neutralizing antibodies that limit the toxin’s usefulness.^291,292,293 A distinction should be made between non-neutralizing antibodies, which may have no clinical significance, and neutralizing antibodies.²⁹⁴

DEPOLARIZING ANTAGONISTS. Extraocular Muscle Effects. The depolarizing antagonists, such as succinyldicholine, decamethonium, and hexacarbacholine (Fig. 2), paralyze striated voluntary muscles throughout the human body but have a different effect on the extraocular muscles. The extraocular muscles respond with a sustained contraction. This was first noted by Hofmann and Holzer,²⁹⁵ who reported that the contraction of the extraocular muscles caused an increase in human intraocular pressure. This observation was supported by Lincoff and co-workers.^296,297 The maximum intraocular pressure reported is 55 mmHg.²⁹⁸

The shortening effect and increased muscle tension created by succinyldicholine have been measured in humans.²⁹⁹ The drug always produced contraction in the horizontal and vertical recti muscles. The results in the oblique muscles were variable.

The explanation is that there are two types of extraocular muscle fibers. In the center of each muscle, typical striated muscle fibers, with a single myoneural junction (en plaque fibers), are most numerous. These are paralyzed by succinyldicholine. However, the surface layers contain fibers with multiple myoneural junctions. These are the “en grappe” fibers. Their innervation has the appearance of a bunch of grapes. It is these fibers that respond to succinyldicholine with a sustained contraction.^300,301,302 The strength of contraction depends on the dose of succinyldicholine and the number of multiply innervated fibers. In sheep superior oblique muscle, succinyldicholine produces only 7% of the force generated by maximum electrical stimulation of the entire muscle.³⁰³ In cats, up to one third of the whole muscle force has been generated by succinyldicholine.³⁰⁴ Succinyldicholine causes a partial reduction in the resting membrane potential and a localized contraction in the area around each myoneural junction.³⁰⁵ Because these junctions are found along the entire length of the en grappe fiber, most of each fiber is depolarized and shortened.³⁰⁶ However, unlike en plaque fibers, which are capable of generating action potentials, the en grappe fibers are believed to respond to both neuronal impulses and succinyldicholine with slow, graded contractions (i.e., they are unable to conduct action potentials).³⁰⁷ This may explain why Kornbleuth and colleagues²²⁹ found a dissociation between ocular muscle electrical activity and the elevation of intraocular pressure. They performed simultaneous tonography and electromyography on subjects given succinyldicholine. They expected to find that the increase in intraocular pressure was proportional to the degree of extraocular muscle electrical activity. Electromyography was performed on antagonistic vertical or horizontal recti of the same eye in subjects receiving 5, 10, 15, or 20 mg succinyldicholine. These doses were too small to produce apnea. A 2- to 4-mmHg increase in intraocular pressure occurred in patients under general anesthesia, and a 5- to 14-mmHg increase occurred in conscious volunteers. The duration, but not the magnitude, of the ocular pressure elevation appeared to be dose related. The rise in intraocular pressure elevation occurred just as the electrical activity of the extraocular muscle was reduced. The inability of the en grappe fibers to conduct action potentials may explain this dissociation.

Curare, but not atropine, blocked the sustained contraction produced by succinyldicholine in animal studies.³⁰⁸ Katz and Eakins²³⁷ found evidence that the en plaque fiber response to depolarizing antagonists was quantitatively different for eye muscles and skeletal muscles (e.g., the dose of succinyldicholine needed to depress the twitch response of the superior rectus muscle was greater than that needed to block skeletal muscles). This difference was less when decamethonium was used.

Each of the six human extraocular muscles contains multiply innervated and singly innervated fibers in about the same proportions: in the periphery, 92% ± 10% are multiply innervated, and in the center, 33% ± 12% are multiply innervated.³⁰⁹ The contractile effect of succinyldicholine on the en grappe fibers is generally considered undesirable. However, Hannington-Kiff³¹⁰ suggested that the anesthesiologist use the resultant change in interlimbal distance to monitor the action of succinyldicholine. The interlimbal distance in 35 patients under general anesthesia before succinyldicholine injection was 52.9 mm ± 4.3 mm. Ninety seconds after injection, this distance had decreased to 49 mm ± 5.5 mm. One minute after the return of spontaneous respirations, the interlimbal distance was near baseline values, 51.4 mm ± 3.3 mm. The duration of succinyldicholine stimulation of en grappe fibers and the duration of succinyldicholine paralysis of en plaque fibers appeared similar. Taylor and associates³¹¹ also found that the intraocular pressure returned to baseline with the onset of spontaneous respirations. Pandey and co-workers³¹² studied the time course of the ocular pressure elevation in 34 patients given succinyldicholine in a dose of 1.5 to 2 mg/kg. An increase in intraocular pressure was present in less than 1 minute, peaked in 2 to 4 minutes, and returned to baseline in 6 minutes.

Succinyldicholine stimulation of the extraocular muscles results in a variable amount of enophthalmos; as much as 3.25 mm has been reported.³¹³ The conjunctival vessels dilate,³¹⁴ and the forced-duction test may suggest a mechanical restriction for up to 20 minutes.³¹⁵

Mindel and colleagues³¹⁶ theorized that the basic deviation of the eyes was produced by the balance of forces exerted by the multiply innervated muscle fibers. They used intravenous succinyldicholine to stimulate these fibers and to remove the influence of the singly innervated fibers, which were paralyzed. Under general anesthesia, esotropic patients became esotropic when 2 mg/kg was injected. Exotropic patients became exotropic.³¹⁷ Despite this qualitative success, the measured amount of drug-induced ocular deviation did not agree closely with that in the conscious patient. These authors believed that this incongruity was due to A and V pattern effects, that is, in conscious patients the eyes could be maintained in the primary position by fixation, whereas in anesthetized patients the eyes could move vertically as well as horizontally. Lingua and associates³¹⁸ have reported similar findings. In addition, they found a correlation between the ocular position induced by succinyldicholine immediately after strabismus surgery and the 1-week postoperative result.

Intraocular Muscle Effects. Abramson³¹⁹ used ultrasound to measure anterior chamber depth and lens thickness in human subjects given 1 mg/kg succinyldicholine intravenously. Succinyldicholine caused a rapid increase in anterior chamber depth and a decrease in lens thickness. This effect was present within 20 seconds, was maximal in 45 to 210 seconds, and disappeared in 3 to 5 minutes. Abramson attributed this effect to a relaxation of accommodation. However, there was no change in the diameter of the pupil. It is unlikely that succinyldicholine would decrease the activity of only one of the two parasympathetic structures innervated by the ciliary ganglion.

Side Effects. Succinyldicholine may produce a bradycardia. This is usually associated with multiple injections. However, asystole has been reported after a single dose.³²⁰ Succinyldicholine and, to a lesser degree, its first hydrolysis product, succinylmonocholine, are responsible for this cardiac effect.³²¹

Succinyldicholine is inactivated by esterase hydrolysis. The apnea produced by respiratory muscle paralysis may be prolonged in patients with genetically³²² or acquired reduced serum cholinesterase activities.^323,324 Chronic systemic absorption from topical cholinesterase inhibitor eyedrops can result in such an effect.³²⁵ In one report, a reduction of serum cholinesterase activity 62% below normal from echothiophate eyedrops permitted endotracheal intubation using a total of only 9.5 mg succinyldicholine.³²⁶ Decamethonium, because it is excreted unmetabolized, is not dependent on the level of serum cholinesterase activity for termination of its effect.

There are at least four alleles, at a single locus, that control the type of serum cholinesterase made by the liver: normal, dibucaine resistant, fluoride resistant, and silent genes. Approximately 1 in 4000 in the population will develop prolonged apnea after succinyldicholine injection. This is the incidence of a person having both genes abnormal. Approximately 1 in 400 in the population will be a normal dibucaine-resistant heterozygote and will develop a shorter but still significantly prolonged apnea. Other factors reducing serum cholinesterase activity are hepatic disease,³²⁷ malnutrition, anemia, severe dehydration, late pregnancy,³²⁸ and systemic absorption of cholinesterase inhibitors. In one study, 5% of a general surgical population had decreased serum cholinesterase activity.³²⁹

The sustained contraction of the extraocular muscles after succinyldicholine injection poses a threat if there is a preexisting perforating laceration or if an incision is to be made into the eye (e.g., cataract extraction). Expulsion of the intraocular contents can result from the steady squeezing of the eye muscles. Lincoff and colleagues²⁹⁷ mentioned three personal communications linking the use of succinyldicholine to this complication. There are several ways it can be avoided: After succinyldicholine injection, the intraocular pressure is monitored with a tonometer until it returns to normal. Use of depolarizing nicotinic antagonists is avoided. Alternatively, a pharmacologic antagonist can be used to prevent the effect. With regard to this last suggestion, cholinesterase inhibitors are not of value. Although they reverse the action of nondepolarizing muscle relaxants (e.g., curare), they prolong and enhance the action of depolarizing drugs.³³⁰ Theoretically, pretreatment with small doses of curare or succinyldicholine itself might prevent succinyldicholine-induced ocular muscle contraction. However, the results are conflicting.^331,332 Macri and Grimes³³³ found that prior curare injection prevented ocular hypertension in cats given succinyldicholine. Because both drugs competed for the same receptor, multiple doses of succinyldicholine overcame the curare effect. Miller and associates³³⁴ found that 1 mg/kg succinyldicholine produced a mean intraocular pressure increase of 8.5 mmHg in 10 control subjects. Parenteral gallamine, 20 mg, or curare, 3 mg, administered 3 minutes before succinyldicholine, prevented this effect in both glaucomatous and normal subjects. However, when Meyers and co-workers³³⁵ studied the value of curare, 0.09 mg/kg, or gallamine, 0.3 mg/kg, given 3 minutes before 1 to 1.5 mg/kg succinyldicholine, they found that ocular pressure elevations still occurred (e.g., in 10 patients with a mean intraocular pressure of 13 ± 1 mmHg, curare did not prevent an elevation to 24 ± 1.3 mmHg). Pancuronium, 0.14 mg/kg, also was ineffective.³³⁶ Katz and colleagues³³⁷ found that hexafluorenium, which is both a nondepolarizing (curarelike) blocking agent and a plasma cholinesterase inhibitor, in doses of 0.4 mg/kg, prevented the rise in intraocular pressure from succinyldicholine, 0.3 mg/kg. Hexafluorenium was effective only if it was injected before succinyldicholine, not after it.

Perhaps the preceding inconsistencies result from the multiplicity of factors that can affect the intraocular pressure (e.g., depth of anesthesia, serum O₂ and CO₂ levels, venous pressure, and tracheal stimulation). Barbiturates, given to induce anesthesia, can reduce muscle endplate currents.³³⁸ When thiopental was injected 2 minutes before succinyldicholine, 1 mg/kg, there was no significant increase in ocular pressure.³³⁹ Macri and Grimes³³³ disinserted all the extraocular muscles in cats. This reduced, but did not prevent, an increase in intraocular pressure after succinyldicholine. Taylor and associates³¹¹ reported that in 25 of 29 patients, the blood pressures 1 minute after succinyldicholine injection were elevated. These may have contributed to the short-term elevations in ocular pressure.

The blood pressure elevations could be because succinyldicholine raises blood catecholamine levels. Succinyldicholine, but not curare, increased plasma norepinephrine levels immediately after injection.³⁴⁰ Succinyldicholine, 1 mg/kg, produced a peak norepinephrine elevation 3 minutes after injection, raising levels from a mean of 301 pg/mL at baseline to 647 pg/mL. The increase had disappeared by 10 minutes. The elevation in epinephrine levels was less marked and became insignificant by 2 minutes.

Wynands and Crowell³⁴¹ found that 50% of patients given succinyldicholine had a rise in intraocular pressure. In 3 of 18 subjects, the elevation was more than 10 mmHg. However, endotracheal intubation alone produced a rise of more than 10 mmHg in 10 of 23 subjects. Lewallen and Hicks³⁴² and Craythorne and co-workers²⁹⁸ reported conflicting results in subjects without intubation. The former found that injection of 40 mg succinyldicholine did not raise human intraocular pressure; the latter found that continuous infusion of 0.1% to 0.2% succinyldicholine did.

A most intriguing study was that of 15 patients having uniocular enucleations.³⁴³ These subjects were anesthetized with 3 to 4 mg/kg thiopental intravenously and maintained with halothane or isoflurane and nitrous oxide/oxygen. No premedications were used. All six extraocular muscles were severed from the eye to be enucleated. Intraocular pressures were then obtained bilaterally. Succinyldicholine 1.5 mg/kg was administered and the intraocular pressures were measured every 30 seconds for 5 minutes. The systolic and diastolic blood pressures were not significantly different before and 90 seconds after succinyldicholine administration, but the intraocular pressures were significantly and maximally elevated bilaterally at this time. Surprisingly, there was no significant difference in the mean ± SE intraocular pressure rise between those eyes with severed muscles and those eyes with muscle insertions intact. The presuccinyldicholine injection intraocular pressures of the normal and muscle-severed eyes were, respectively, 15.1 ± 1 mmHg and 16.1 ± 1 mmHg. At 90 seconds after succinyldicholine injection, the respective intraocular pressures were 25.2 ± 1.6 mmHg and 24.7 ± 1.8 mmHg. Perhaps an increase in venous blood pressure produced these bilateral results, but the apparent lack of an extraocular muscle effect is inexplicable.

GENERAL CONSIDERATIONS. The ocular effects of this class of drugs include reduction in intraocular pressure; stimulation of the iris sphincter, producing miosis; and stimulation of the ciliary muscle, producing accommodation. Examples of direct-acting muscarinic drugs are muscarine, pilocarpine, aceclidine (3-acetoxyquinuclidine), arecoline, and acetyl β-methylcholine (methacholine, Mecholyl) (Figs. 3, 4, 5). In high concentration, pilocarpine also stimulates choline acetyltransferase synthesis of acetylcholine,³⁴⁴ but this is probably of little clinical significance.³⁴⁵

Isopilocarpine is a naturally occurring stereoisomer of pilocarpine present in very small amounts in most preparations of the drug.³⁴⁶ Isopilocarpine has one tenth the binding affinity of pilocarpine for bovine ciliary muscle and has very little pharmacologic activity.

Carbachol (carbamylcholine) is both a direct-acting muscarinic agonist and a direct-acting nicotinic agonist. In addition, it has indirect agonist activities (i.e., it is a cholinesterase inhibitor). This last action results from the NH₂ substitution on the acetyl group of carbachol. It markedly reduces the molecule’s susceptibility to hydrolysis by acetylcholinesterase but does not prevent carbachol from competing with acetylcholine for the anionic and esteradic sites of acetylcholinesterase. This interference with acetylcholine hydrolysis results in a further increase in cholinergic activity. Aceclidine, too, is slightly cholinesterase resistant and has weak anticholinesterase activity.

Pilocarpine’s structure is quite different from that of acetylcholine. Either of the nitrogen atoms in the imidazole ring can become positively charged and bind to the anionic receptor site. Pilocarpine does not have the classic muscarinic agonist structure of acetylcholine (N-C-C-O-C-C). However, pilocarpine’s interatomic spacing (N-C-C-C-C) may mimic it.³⁴⁷ Evidence exists that the intrinsic muscarinic activity of pilocarpine is less than that of acetylcholine and carbachol.^348,349 However, additional factors play a role in determining clinical efficacy. These are primarily pharmacokinetic. For example, acetylcholine and acetyl β-methylcholine are hydrolyzed by tissue esterases and may not reach the site at which their action is desired. Inhibiting tissue cholinesterases increases the efficacy of both drugs. Arecoline, aceclidine, and pilocarpine are tertiary amines and can penetrate the corneal epithelium more readily than the positively charged quaternary amines such as acetylcholine, carbachol, and acetyl β-methylcholine. This correlates with the lipid solubilities of these drugs. As measured with toluene: pH 6.5 buffer, the partition coefficient of arecoline is 0.1; pilocarpine, 0.033; aceclidine, 0.002; and carbachol, <0.0001.³⁵⁰ The pHs of eyedrops and tears alter the proportions of nonionized drug (e.g., the ability of arecoline and oxotremorine to reverse mydriasis increases as a function of the pHs of their solutions).³⁵¹ Drugs that are ionized at all pHs, such as carbachol, are concentration dependent, not pH dependent. The poor corneal penetration of carbachol resulted in Clarke³⁵² finding marked variability in its ability to lower intraocular pressure. O’Brien and Swan³⁵³ performed a corneal massage through the lids of human subjects. This allowed a 0.09% carbachol solution to be as effective a miotic as a 1.5% to 4.5% carbachol solution. Topical anesthetics and surfactants also aided penetration (e.g., the addition of 0.03% benzalkonium to 1.5% carbachol produced significant accommodation for 48 hours).

Accurate estimates of corneal penetration by pilocarpine have been impaired by the use of tritiated molecules. Asseff and co-workers³⁵⁴ gave anesthetized monkeys eyedrops of 1% to 8% tritiated pilocarpine. They reported approximately 3% corneal penetration at 5 minutes. Chrai and Robinson,³⁵⁵ studying rabbits, found similar levels after applying unpurified tritiated pilocarpine but only 0.13% corneal penetration at 5 minutes after applying purified material. Maximum penetration occurred at 20 minutes and was 0.18%. At 2 hours, the aqueous humor concentration was 0.03% of the administered dose. These authors attributed the previous higher values to tritium exchange between pilocarpine and its solvent during storage. Penetration of the tritiated solvent resulted in falsely high estimates of corneal penetration. Krohn and Breitfeller³⁵⁶ applied two drops of 2% pilocarpine HCl (approximately 2000 μg) to corneas of patients under general anesthesia; the pilocarpine concentration in the aqueous humor was not more than 5 μg/mL.

Pilocarpine can be administered from continuous-release membranes (Ocusert) made of polymerized ethylenevinylacetate. Ideally, the low-dosage form (Ocusert P20) releases 20 μg/h and the high-dosage form (Ocusert P40) releases 40 μg/h. However, Armaly and Rao³⁵⁷ found that the in vitro release rates from membranes supposedly dispensing 20 μg/h, 50 μg/h, and 80 μg/h were two and one half to three times their projected rates during the first 7 hours of use. Between 7 and 24 hours, the release rates remained slightly elevated, but within the next 24 hours they fell to slightly below their projected levels and continued to decline. After 3 to 4 days, the release rates were 78%, 88%, and 81% of those expected. These results may explain why the glaucoma of two of Armaly and Rao’s patients was controlled for the first 2 days of membrane use but not thereafter.

Pilocarpine has also been applied as a polymer emulsion^358,359 and as a gel. Both provide prolonged drug delivery to the eye and, compared with drops, longer duration of action. The gel is available commercially as 4% pilocarpine HCl in a viscous aqueous acrylic gel vehicle; this concentration of pilocarpine approaches the binding limit of the polymer used to make the gel. A single daily application of the gel has a hypotensive effect for 24 hours. For the first 18 hours, this effect is indistinguishable from that elicited by 4% pilocarpine eyedrops given four times a day in terms of both pressure and pupillary diameter.³⁶⁰ For hours 18 to 24, the drops are more effective.³⁶¹

Pathology can alter pharmacokinetics and drug response. Patients with such diverse pathology as atopy and trisomy 21 are said to be more sensitive to muscarinic agonists and antagonists.^362,363

Patients with intraocular inflammation inactivate pilocarpine. Schonberg and Ellis³⁶⁴ found that the primary aqueous humor of humans and rabbits did not inactivate pilocarpine, but serum and secondary aqueous did. Ten percent of 500-μg pilocarpine was inactivated by 200 μL of secondary aqueous humor in 1 hour. Inactivation could be prevented by heating serum to 60°C for 10 hours, by dialysis, or by the addition of some (e.g., penicillamine and ethylenediaminetetraacetic acid), but not all, chelating agents. Inactivation could not be inhibited by anticholinesterase drugs. Rabbit serum was more active against pilocarpine than human serum. Only at high pilocarpine concentrations were human ocular tissues (cornea, iris-ciliary body, lens, retina, and choroid) more active than the corresponding rabbit ocular tissues.³⁶⁵ Lee and colleagues³⁶⁶ found that pigmented rabbit corneas inactivated pilocarpine much more effectively than did albino rabbit corneas.

OCULAR PHARMACOLOGY Iris Sphincter Muscle. ANIMAL STUDIES. Pilocarpine has less intrinsic muscarinic activity than the endogenous transmitter acetylcholine. As a result, Swan and Gehrsitz³⁶⁷ found that rabbits given topical 0.25% physostigmine or 0.1% DFP had less miosis if 4% pilocarpine was previously administered. If pilocarpine were injected into the cornea after maximum DFP miosis, a transient dilatation occurred. These results were consistent with pilocarpine-acetylcholine competition for the same receptors.

Miosis in mammals is associated with M₃ receptor activation and increased phospholipase C activity.^368,369 Feedback inhibitory prejunctional muscarinic receptors on sympathetic neurons of the iris dilator muscle seem to be of a different subtype.¹⁹⁹ In rabbits, these prejunctional receptors respond to methacholine, oxotremorine, muscarine, and carbachol by inhibiting norepinephrine release; atropine can block this effect.³⁷⁰

Denervation supersensitivity of the rat iris sphincter is not associated with a change in muscle fiber resting potential but is associated with increased sensitivity to calcium.^371,372 Thus, supersensitivity could be due to an increased release of intracellularly bound calcium and/or an increased influx of extracellular calcium.

Bito and Banks³⁷³ administered DFP drops chronically to monkeys and then discontinued the drug. They observed several interesting pupillary phenomena, one of which was a diminished response to direct-acting muscarinic agonists during the recovery period. For at least 3 days after discontinuing DFP, the pupils responded to light but did not respond to carbachol or pilocarpine. Similar results were obtained in the cat.³⁷⁴ These results may have been produced from an acquired subsensitivity of the sphincter muscarinic receptors (i.e., a subsensitivity produced by the prior high acetylcholine concentrations). Another possibility is that the greater intrinsic agonist activity of acetylcholine, compared with that of pilocarpine and carbachol, became manifest. Other data from this laboratory,³⁷⁵ confirmed by Claesson and Barany,³⁷⁶ suggested that an alteration of muscarinic receptor sensitivity had occurred. Exposure of feline eyes in vivo to continuous light resulted in subsensitivity to pilocarpine, whereas light deprivation resulted in increased sensitivity. After 7 days in ambient light of 1200 lumens/m², the irides became insensitive to pilocarpine. Surprisingly, subsensitivity to carbachol or acetyl β-methylcholine could not be demonstrated in vitro. Hemicholinium (which reduces acetylcholine synthesis) injected intravitreally in vivo resulted in increased sensitivity to administered muscarinic agonists. Barany³⁷⁷ reported that chronic administration of pilocarpine drops did not produce subsensitivity in experimental animals challenged with intravenous pilocarpine. However, after prolonged treatment in the form of continuous-release membranes, pilocarpine did produce subsensitivity.

Perhaps the explanation for the aforementioned phenomena is miosis from peptide transmitters. There is evidence that the neurotransmitter substance P, identified in trigeminal nerve fibers, produces miosis.³⁷⁸ Mandahl³⁷⁹ has suggested that, in the rabbit, the miosis elicited by cholinesterase inhibitors and pilocarpine is primarily the result of release of substance P from trigeminal nerve endings in the iris. According to this theory, continuous application of cholinesterase inhibitors and pilocarpine would deplete the stores of substance P, resulting in a reduced response, whereas the acetylcholine release by light-stimulated parasympathetic fibers would remain intact.

HUMAN STUDIES. Lowenstein and Loewenfeld³⁸⁰ and Newsome and Loewenfeld³⁸¹ studied the iris using pupillography. Drops of 1% to 2% pilocarpine were given to volunteers without ocular disease. The onset of miosis, maximum miosis, and duration of miosis varied greatly from person to person. Maximum miosis occurred in 25 to 45 minutes, was maintained for 1 to 3 hours, and required 24 to 36 hours for full recovery. The pupillary reflex to light persisted unless miosis was maximal. However, during the recovery phase, the light response was markedly diminished. When submaximally effective concentrations of pilocarpine (e.g., 0.1%) were followed by submaximal concentrations of a cholinesterase inhibitor (e.g., 0.1% physostigmine), the results were additive.

Ogle and associates³⁸² and Morgan and co-workers³⁸³ also used pupillography to study miosis in normal subjects. They reported latencies after 1% pilocarpine of 9.8 ± 0.9 minutes. The latencies for 0.75% carbachol and 0.25% physostigmine were 13.7 ± 2.2 minutes and 13.9 ± 1 minutes, respectively. The rates of miosis and amounts of maximum miosis were less for carbachol than for pilocarpine and physostigmine, yet all three drugs required about the same time to produce their maximum effect. The times required for recovery to 50% of the predrop pupil diameters were: carbachol, 164 ± 46 minutes; pilocarpine, 271 ± 109 minutes; and physostigmine, 456 ± 28 minutes. The latency of mydriasis from 0.5% tropicamide (Mydriacyl) was similar to the latency of miosis from pilocarpine. The changes in pupillary diameter due to light stimuli during the latency periods of both pilocarpine and tropicamide were similar; the amounts and maximum rates of pupil constriction after onset of action also were similar.

After the short-term application of various direct-acting muscarinic agonists, the response produced by the light reflex was reduced.³⁸⁴ This may have represented competition between residual exogenous agonists, with weaker intrinsic activity, and the endogenous agonist acetylcholine, with more potent intrinsic activity. The light reflex required approximately 24 hours to recover after 2 drops of 2% pilocarpine, 7 hours to recover after 10 drops of 0.2% arecoline hydrobromide, and 24 hours to recover after 10 drops of 0.2% aceclidine hydrochloride.

Ciliary Muscle. ANIMAL STUDIES. Tornqvist^385,386,387 studied accommodation in monkey eyes. Intramuscular pilocarpine over a dose range of 0.2 to 2 mg/kg appeared to have equal affinity for iris and ciliary muscle. The lowest dose caused 2 diopters (D) or less of accommodation, and the highest dose always caused more than 12 D of accommodation. One monkey was given higher doses of pilocarpine, and the myopia paradoxically decreased: 2 mg/kg produced 20.3 D of accommodation, 5 mg/kg produced 11.7 D of accommodation, and 50 mg/kg produced 8 D of accommodation. Similarly, small doses of systemic carbachol, 0.005 to 0.1 mg/kg, produced increasing accommodation, but larger doses, 0.1 to 0.5 mg/kg, produced decreased accommodation. This phenomenon could not be reproduced by using high concentrations of topical muscarinic agonists. Retrobulbar anesthesia did not prevent this paradoxical effect, indicating that central nervous system and ganglionic effects were not the cause. Intravenous atropine prevented, or reversed, the paradoxical effect. A topical dose of pilocarpine that produced half the maximum accommodative effort was 100 times more than that needed to produce half the maximum miosis. Because systemically administered pilocarpine did not exhibit this difference, it was concluded that pharmacokinetic factors resulted in more drug reaching the iris than reached the ciliary muscle after topical application.

Barany³⁸⁸ studied subsensitivity of the monkey’s ciliary muscle after a single drop of topical carbachol given unilaterally. For approximately 1 week, the accommodative response in the treated eye was less than that in the untreated eye after systemic administration of pilocarpine, carbachol, or bethanechol. The reasons for this effect were not clear, but the author did not believe the answer was a reduction in the number of muscarinic receptors or a reduction in their binding characteristics.

In monkeys, the loss of accommodation that occurs with aging is not associated with a reduction in the number of ciliary muscle muscarinic receptors.³⁸⁹

The agonist aceclidine is less effective in producing accommodation than in increasing miosis and outflow facility. Studies in monkey eyes found that M₃ receptors predominate in ciliary muscle, iris sphincter muscle and trabecular meshwork (i.e., the differences in pharmacologic responses could not be attributed to different muscarinic receptor subtypes).^390,391

After ciliary ganglionectomy, monkey ciliary muscle supersensitivity to pilocarpine was associated with a 12% to 84% reduction in muscarinic receptors.³⁹² The magnitude of this seemingly paradoxical finding is difficult to explain simply by attributing it to the loss of prejunctional muscarinic receptors on nerve endings.

HUMAN STUDIES. Ciliary body thickness was measured in volunteers by the use of ultrasound.³⁹³ Two hours after applying a drop of 2% pilocarpine, the increase in mean thickness was 0.06 mm. Anterior chamber depth was measured after single and multiple drops of 2% or 4% pilocarpine.³⁹⁴ All subjects showed a decrease in anterior chamber depth after a single drop of pilocarpine. This shallowing remained significant for the 6 hours that measurements were made. When compared with the untreated eye, a single drop of 2% pilocarpine produced a mean maximum shallowing of 0.09 mm that occurred 3 hours after instillation; 4% pilocarpine produced a mean maximum shallowing of 0.07 mm that occurred 4.5 hours after instillation. Pilocarpine (2%), every 6 hours, produced a mean maximum anterior chamber narrowing of 0.12 mm. This was not significantly different from the single-drop effect. One week after chronic therapy was discontinued, the mean depth of the treated eye was not significantly different from that of the control eye. Abramson and colleagues³⁹⁵ confirmed that the anterior chamber shallowing effect from 2% pilocarpine, as well as the increase in lens thickness, was similar after single-drop and chronic (10-day) therapy.

Hallden and Henricsson³⁹⁶ produced with-the-rule astigmatism in 10 young men by asymmetrically contracting the ciliary body. This was achieved by applying a cotton pledget containing 10% pilocarpine to the temporal bulbar conjunctiva for 3 minutes, creating 0.75- to 2 D myopia with an axis between 0° and 20°.

Intraocular Pressure. The site at which muscarinic drugs act to lower intraocular pressure is a subject of controversy. Their effect is believed to be local, although an occasional report raises the possibility of a systemic effect (e.g., Willetts³⁹⁷ reported that normotensive subjects given unilateral pilocarpine drops had a bilateral decrease in ocular pressure). However, subjects with a mean plasma level of 2.9 ng/mL, delivered by skin patches, did not have a significant reduction in intraocular pressure.³⁹⁸

The different mechanisms of action that have been proposed include a decrease in aqueous humor secretion; an increase in removal of aqueous humor by active transport; an increase in removal of aqueous humor by passive trabecular outflow (from increased iris sphincter tone, increased ciliary muscle tone, or decreased episcleral venous pressure); and an increase in removal of aqueous humor by way of nontrabecular routes.

AQUEOUS HUMOR SECRETION. The data from different laboratories are inconsistent. Becker³⁹⁹ found that pilocarpine drops increased rabbit aqueous humor formation in vivo. Green and colleagues⁴⁰⁰ reported that acetylcholine and pilocarpine produced an increase in rabbit aqueous humor secretion in vitro. Macri and Cevario⁴⁰¹ administered pilocarpine intra-arterially to enucleated cat eyes and also found an increase in aqueous humor secretion; however, outflow facility was increased, producing an overall reduction in intraocular pressure. Berggren^402,403 found the opposite results. He used in vitro thinning of rabbit ciliary processes as a measure of secretion. Pilocarpine 10^-9 mol/L, physostigmine 10^-7 mol/L, acetylcholine 10^-5 mol/L, and carbachol 10^-5 mol/L inhibited secretion. The therapeutically inactive stereoisomer of pilocarpine, isopilocarpine, did not block secretion. Atropine 10^-7 mol/L alone had an inhibitory effect⁴⁰² and also prevented the more potent pilocarpine effect.⁴⁰³ However, when intravenous pilocarpine, 2 mg/kg, was injected in vivo 5 minutes before the in vitro studies, there was no evidence of altered secretion.

Edwards and colleagues,⁴⁰⁴ using constant-pressure perfusion, measured the effect of unilateral topical 4% pilocarpine on monkeys under general anesthesia. Pilocarpine caused an increase in aqueous humor production. However, Walinder and Bill⁴⁰⁵ found in vervet monkeys that infusion of pilocarpine 10^-4 mol/L into the anterior chamber produced a reduction in aqueous humor formation of approximately 1 μL/min; atropine infusion 3 × 10^-5 mol/L abolished the pilocarpine effect.

Barsam⁴⁰⁶ found that the hypotensive effect of pilocarpine lasted longer than the duration of increased outflow facility in human subjects. He believed that pilocarpine was producing a reduction in aqueous humor secretion. However, fluorophotometry of normotensive eyes has provided evidence that pilocarpine has only a minor effect on aqueous humor formation, which is to increase secretion by approximately 14%.⁴⁰⁷ This may or may not be true for the glaucomatous eye as well.

Muscarinic agonists might alter aqueous humor secretion by altering ciliary blood flow. Stjernschantz and Bill⁴⁰⁸ studied cholinergic control of uveal blood flow in anesthetized animals. Stimulation of the intracranial third nerve altered the blood flow to the iris and ciliary body but not to the retina and choroid. In the cat, the ciliary body blood flow was increased; in the rabbit, it was decreased; and in the monkey there was no statistically significant effect. In all three species, the iris blood flow was decreased by third-nerve stimulation. Intravenous atropine blocked this iris effect in all three species and prevented the increased ciliary body blood flow in cats.

AQUEOUS HUMOR ACTIVE TRANSPORT. Large vacuoles have been observed in the endothelium of the inner wall of Schlemm’s canal. It has been suggested that they represent the result of active transport of aqueous humor. Holmberg and Barany⁴⁰⁹ found that monkey eyes treated with pilocarpine had a reduced number of vacuoles. Lutjen-Drecoll⁴¹⁰ placed 20 μg pilocarpine hydrochloride into the anterior chambers of monkeys and then studied the eyes with electron microscopy. Large vacuoles were rarely encountered. Grierson and associates⁴¹¹ studied the enucleated eyes of eight patients with melanomas. Pilocarpine drops were given before their removal. These were compared with 11 control eyes but not in a masked manner. There were more than twice the number of large vacuoles in the pilocarpine-treated eyes. However, the authors’ results from baboon eyes led them to conclude that the increased vacuolization was not due to a direct effect of pilocarpine on the endothelium of Schlemm’s canal. Instead, they attributed the vacuoles to a secondary effect from increased trabecular flow.⁴¹²

INCREASE IN PASSIVE AQUEOUS HUMOR OUTFLOW. Most investigations have dealt with this mechanism of muscarinic action. Edwards and co-workers⁴⁰⁴ found that topical 4% pilocarpine caused more than a doubling in monkey eye outflow facility, from 0.41 ± 0.04 μL/min/mmHg to 1.09 ± 0.14 μL/min/mmHg. Atropine alone had no effect. Barany⁴¹³ found that pilocarpine, administered topically or injected into the anterior chamber, increased vervet monkey facility of outflow. Two components appeared to be present, because atropine, injected intravenously or intraocularly, reversed part of the increase in 3 to 5 minutes and the remainder in approximately 30 minutes. Lutjen-Drecoll⁴¹⁰ studied electron micrographs of monkey eyes and found that the trabecular pore area in contact with the inner wall of Schlemm’s canal correlated, after pilocarpine administration, with the outflow facility.

In humans, the hypotensive effect of pilocarpine is usually, but not always, associated with an increase in outflow facility. Outflow facility, accommodation, and pilocarpine-induced accommodation decrease with age. However, the pilocarpine-induced increase in outflow facility and reduction in intraocular pressure were found not to decrease with age.⁴¹⁴ Pilocarpine, using gonioscopy to evaluate the drug’s effect, seems to produce a bowing inward of the trabecular meshwork toward the anterior chamber and away from the canal of Schlemm.⁴¹⁵