Nictitating Membranes Research Articles

The influence of the nictitating membrane on steroid inhibition of limbal wound healing.

In a study involving 34 rabbits, it was found that local corticosteroids inhibit limbal wound healing in normal rabbit eyes, and do not inhibit similar wounds if the nictitating membrane has been removed. It was further found that the contact time of fluorescein labelled corticosteroids is almost 3 times longer in rabbit eyes with nictitating membranes than those in which the membranes have been removed.

Actions of amphetamine and antagonists on pupil diameter in the chronic sympathectomized dog

The left superior cervical ganglia were removed from 5 dogs. Beginning 30 days postoperatively, epinephrine (10 microgram/kg/min), norepinephrine (10 microgram/kg/min), and d-amphetamine (1.0 mg/kg) were infused i.v. for 10 min following either vehicle, phenoxybenzamine, pimozide, or haloperidol. Epinephrine and norepinephrine dilated the pupil and retracted the nictitating membrane of the denervated side, whereas amphetamine dilated both pupils and retracted both nictitating membranes. Phenoxybenzamine (4 mg/kg) constricted primarily the pupil of the innervated iris and completely antagonized the effects of the catecholamines on the irides and amphetamine on the nictitating membranes, but only partially antagonized amphetamine-induced mydriasis. Haloperidol (1.0 mg/kg) constricted both pupils, possessed only modest alpha-adrenergic blocking activity, and was as effective as phenoxybenzamine in antagonizing amphetamine-induced mydriasis. Pimozide (0.1 mg/kg) constricted both pupils, had no significant alpha-adrenergic blocking activity, and did not antagonize amphetamine-induced mydriasis. Pimozide and haloperidol, but not phenoxybenzamine, blocked amphetamine-induced stereotyped head bobbing. These results suggest that amphetamine produces mydriasis in the dog through a peripheral sympathetic action and also through a central mechanism involving inhibition of the oculomotor nucleus. However, the role of dopamine is not clear.

Electron Microscopic Observations on Smooth Muscles of Cat Nicitating Membrane

The cat nictitating membrane is commonly used for physiological and pharmacological study, since its smooth muscles are virtually all innervated by sympathetic nerves. Thus, morphological studies at the fine structural level have been focussed to the innervation of the nictitating membrane, but not to the smooth muscle proper.The nictitating membranes of adult cats were fixed in situ in protruded state with 2% formaldehyde and 2% glutaraldehyde in 0.1 M cacodylate buffer (pH. 7.3). The tissues were then dissected out to continue fixation in the same buffer. After rinse in 10% sucrose in the same buffer, the tissue blocks were postfixed in 1% OsO4 and embedded in Epon.

The Healing of Mechanical Injuries in the Nictitating Membrane of the Frog

Uncomplicated perforating injury of frog nictitating membrane evokes a marked migration of cellular elements.It appears most striking in anterior epithelium but occurs also in the posterior layer. One week after injury the wound is partially filled and complete occlusion occurs 3 to 4 weeks later (at 24 ± 1°C). The injury stimulates DNA synthesis and mitosis in anterior and posterior epithelia. Incorporation of 3H-thymidine has also been observed in the stroma cells. However, the uptake in the posterior epithelium is most pronounced. It becomes conspicuous about 2 days after the infliction of the wound and it remains high for at least one week after that. Stromal cells increase in number. Their origin, as in the case of the cornea, is problematical. The suitability of nictitating membranes for injury studies is discussed.

Adrenergic re-innervation of smooth muscle of nictitating membrane by preganglionic sympathetic fibres.

1. The preganglionic and post-ganglionic trunks of the cervical sympathetic nerve were joined in an end-to-end anastomosis after excision of the superior cervical ganglion in the cat.2. Seventy-five days after the anastomosis the diameter of the pupil was nearly normal and there was almost complete recovery of the prolapsed palpebra and of the nictitating membrane. The contraction of the nictitating membrane, induced by electrical stimulation, caudally to the point of anastomosis, showed that the smooth muscle of the nictitating membrane had been re-innervated.3. Neither hexamethonium nor nicotine had any marked effect on the contraction of the nictitating membrane. Severing the regenerated nerve trunk produced a degeneration contraction. These results are strong evidence that the denervated membranes were re-innervated by true cholinergic preganglionic fibres.4. Our pharmacological studies indicated that in the re-innervated preparations neuromuscular transmission was adrenergic in the sense that it was blocked by phentolamine and not by atropine. These results were confirmed by the histochemical-fluorescence studies which showed that the endings of the regenerated axons contained high concentrations of catecholamines.5. Electron microscopy showed that the regenerated terminals contained none of the small dense-core vesicles, considered to be typical of adrenergic nerve endings, but contained clear synaptic vesicles and an unusually great number of large granular vesicles.6. Our results suggest that the denervated nictitating membranes were re-innervated by cholinergic presynaptic sympathetic fibres that had been modified so that they could release catecholamines in addition to, or instead of, acetylcholine.

Angiotensin-induced contraction of the cat nictitating membrane

We have studied the effect of angiotensin on nictitating membranes of spinal and pentobarbital-anesthetized cats in vivo and in vitro. In all preparations, angiotensin caused dose-related contractions. In vivo, the contractile effect was very weak, rarely more than 5% of maximum, and was demonstrated best by injecting the angiotensin intra-arterially toward the nictitating membrane so that it by passed the ipsilateral superior cervical ganglion. Contractions produced in this way by 0.3–2 μg/kg were not inhibited by removal of the ganglion or the ipsilateral eyeball or by intravenous injection of phentolamine (2mg/kg), atropine (0.5 mg/kg), or gallamine (3 mg/kg). In vitro, angiotensin's contractile effect was stronger, sometimes more than 20% of maximum, but again was not inhibited by phentolamine (3 × 10 -7 g/ml), atropine (3 × 10 -8 g/ml), or methyl atropine (3 × 10 -8 g/ml). Therefore, we conclude that angiotensin can exert a contractile effect upon the nictitating membrane that is independent of both adrenergic and cholinergic mechanisms.

Development of degeneration contraction and supersensitivity in the cat's nictitating membrane after 6-hydroxydopamine.

After subcutaneous injection of 30 mg/kg of 6-hydroxydopamine into the conscious cat the denervated and the decentralized nictitating membranes contracted immediately. While the response of the denervated membrane is of short duration (1.5 h), the decentralized membrane is half relaxed after 11 h. The duration of the latter contraction is similar to that of the degeneration contraction seen after surgical denervation. It is proposed that the response of the decentralized nictitating membrane to 6-hydroxydopamine corresponds to the degeneration contraction. Treatment of the cats with pargyline prolongs both types of degeneration contractions. However, while beta-TMlO {[2-(2,6-dimethylphenoxy)propyl] trimethylammonium chloride had profound effects on the degeneration contraction seen after surgical denervation, it failed to affect that after 6-hydroxydopamine. Indirectly acting sympathomimetic amines prevented the usual effect of beta-TMlO on the degeneration contraction seen after surgical denervation. It is proposed that the failure of beta-TMlO to affect the response of the decentralized nictitating membrane to 6-hydroxydopamine is due to a similar preventive effect of 6-hydroxydopamine. The development of supersensitivity to (-)-noradrenaline was determined at various time intervals after the injection of 6-hydroxydopamine; pithed cats and isolated nictitating membranes were used. Under both conditions the membranes were in a state of sustained contraction, if the experiment was begun before the response of the decentralized nictitating membrane to 6-hydroxydopamine had worn off. After 6-hydroxydopamine the development of supersensitivity coincided with the falling phase of the degeneration contraction. Very similar results had been obtained earlier during the degeneration contraction after surgical denervation; the temporal relation between degeneration contraction and development of supersensitivity is very similar after surgical and chemical denervation.

Supersensitivity of decentralized and denervated nictitating membranes to barium.

SummaryDenervation and decentralization of the nictitating membrane produced a 7-fold increase in sensitivity to barium. The action of barium was not mediated via the release of endogenous norepinephrine. This supersensitivity to barium is consistent with the hypothesis that the postsynaptic form of supersensitivity in smooth muscle is the result of a functional change beyond the level of drug receptors, perhaps in membrane properties.

Supersensitivity of the isolated nictitating membrane of the cat to sympathomimetic amines after impairment of the intraneuronal mechanisms of inactivation

Dose-response curves for (−)-noradrenaline, (−)-adrenaline and (−)-phenylephrine were determined on isolated nictitating membranes after pretreatment with reserpine (to impair vesicular retention of the amines), with pargyline (to block monoamine oxidase, MAO), or with both. Sensitivity was found to be increased by reserpine < pargyline < combined pretreatment. In addition, pretreatment with pargyline increased the slope of dose-response curves as well as the time required for the development of steady-state responses. The degree of supersensitivity to (−)-phenylephrine induced by the combined pretreatment was nearly as great as that produced by denervation. For (−)-noradrenaline and (−)-adrenaline it was smaller than that seen after denervation. Evidence is presented in favour of the view that desensitization to (−)-noradrenaline develops during prolonged exposures to this amine. The pretreatments had no effect on the sensitivity of the denervated membrane to the three amines.

The effect of cocaine on uptake of and sensitivity to noradrenaline in isolated nicitating membranes before and after storage in the cold.

Experiments were carried out on fresh isolated cat nictitating membranes as well as on muscles stored in the cold for 7 days. Storage reduced the cocaine-induced supersensitivity to (−)-noradrenaline but did not abolish it it also reduced responses to tyramine, and about halved the noradrenaline content of the tissue. Cocaine failed to potentiate responses of fresh or of stored muscles to the methoxamine (which is not taken up by adrenergic nerves).

Effects of denervation on the sensitivity of the superior cervical ganglion of the cat to acetylcholine and McN-A-343.

Supersensitivity of the denervated superior cervical ganglion of the cat to acetylcholine (Ach) has been reported by workers who measured contractions of the nictitating membrane, and denied by others who measured postganglionic action potentials as an index of ganglion activity. The present experiments were performed to find out the reason for this discrepancy. The increased responses of nictitating membranes to intra-arterial administration of Ach to denervated ganglia were found to be due to increased sensitivity of membranes to the adrenergic neurotransmitter rather than to an increase in the sensitivity of the ganglia. McN-A-343, a muscarinic agent, produced a greater maximal contraction of the membrane when injected to denervated than when injected to control ganglia. These results suggest that denervation causes an increase in the number and (or) sensitivity of muscarinic receptors in ganglia. Hexamethonium antagonized Ach less effectively on denervated than on control ganglia. The effect of Ach on denervated ganglia was blocked by hexamethonium plus atropine, which suggests that Ach acts both on nicotinic and muscarinic receptors in denervated ganglia. Atropine but not hexamethonium blocked the effect of McN-A-343 on both control and denervated ganglia.

Response of the nictitating membrane to prostaglandin E 1 and angiotensin

Prostaglandin E 1 has previously been shown to potentiate contractile responses of smooth muscle to various stimuli. To study this phenomenon further, responses of cat nictitating membranes to angiotensin and preganglionic electrical stimulation were measured, and so were the changes in these responses following intraarterial administration of PGE 1. In adrenalectomized animals PGE 1 contracted the nictitating membrane independent of the superior cervical ganglion, but response was decreased following adrenergic or cholinergic blockade, and abolished following both. Angiotensin also contracted the nictitating membrane in adrenalectomized animals independent of the superior cervical ganglion, and the response was inhibited by cholinergic but not by adrenergic blockade. PGE 1 potentiated responses to acetylcholine, norepinephrine and angiotensin. It also antagonized ganglionic blockade by hexamethonium.

The nictitating membranes of primates.

AbstractOf 22 species of subhuman primates examined, 21 have a well developed nictitating membrane; it is vestigial only in Pan satyrus/troglodytes. The sizes of membranes range from those that cover only one‐tenth of the eye (Gorilla gorilla) to those that extend over the entire eye (Perodicticus potto and Arctocebus calabarensis). Nictitans, best developed in the lower primates, have a free border that is generally heavily pigmented. With the exception of Galago crassicaudatus, all prosimians studied have a membrane that contains a cartilaginous band; cartilage is also present in Cacajao rubicundus, Cebus albifrons, and Saimiri sciureus. The outer surface of the nictitans of all species of anthropoids has mucous glands; that of the prosimians has few of them. Serous glands are present only in the Lorisoidea; in anthropoids serous glands are confined to the adjacent conjunctiva. Mast cells occur with great frequency in prosimians. Elastic fibers are better developed in anthropoids. Muscle fibers are present only in the nictitating membranes of Lemur monoz and Callithrix aurita. Vellus hairs grow on the outer surface of the nictitans in many of the species studied.

Mechanisms of the pressor and depressor actions of St 155 (2-(2,6-dichlorophenylamino)-2-imidazoline hydrochloride, catapres ®)

St 155 (Catapres ®) has recently been found effective in the treatment of hypertension. The mechanism of its antihypertensive action has been investigated in animal experiments. These indicate that it differs from other clinically useful antihypertensives in its mode of action. In the cat, intravenous injections of St 155 produced an initial rise in blood pressure and a contraction of the nictitating membranes. After treatment with cocaine the pressor response was unchanged, and the contraction of the nictitating membrane was not reduced after denervation of the membrane. These sympathomimetic effects of St 155 are therefore not due to the release of endogenous catecholamines. Since these effects were blocked by phentolamine, they appear to be due to an action on the α-adrenoreceptors. Following the initial pressor response, St 155 produced a long lasting reduction of the blood pressure. St 155 had no ganglion blocking or adrenergic neurone blocking activity and did not block α- or β-adrenoreceptors; the depressor response was therefore not due to impairment of peripheral sympathetic function. The fall in blood pressure was unaffected by pretreatment with atropine or by propranolol, and was therefore not due to effects on muscarinic receptors or β-adrenoreceptors. However there was no fall in blood pressure in spinal cats or in cats treated with guanethidine. It is suggested that the depressor action of St 155 may therefore be due to a central reduction of sympathetic outflow to the heart and blood vessels.

Adrenergic nerve function, noradrenaline level and noradrenaline uptake in cat nictitating membrane after reserpine treatment.

AbstractAfter reserpine injection to cats (0.1 mg/kg subcutaneously) there was a severe impairment of the adrenergic transmission in the nictitating membranes with recovery after 48 to 72 hrs. The noradrenaline of the nictitating membrane was virtually completely lost at the time of the disturbed nerve function and showed no significant restoration at the time of functional recovery. The uptake of tritiated noradrenaline by the nictitating membranes was blocked by reserpine but was partially restored after 48 to 72 hrs, thus coinciding with functional recovery. It is suggested that an intact adrenergic transmission is dependent on the ability of the amine storage granules to incorporate noradrenaline in a small fraction immediately important for the function.

Transplantation of the trematode Philophthalmus hegeneri to new and altered sites

Six of 10 juvenile eye flukes, Philophthalmus hegeneri Penner and Fried 1963 homotransplanted into chicks' eyes survived, grew, and developed. Two flukes that developed in the same eye of a chick appeared fertile. The other 4 flukes developed individually in chicks' eyes and were infertile. The absence of live flukes implanted into the coeloms of chicks indicated an incompatibility of this site. Flukes implanted into the cloacas of chicks showed no signs of growth or development and were dead by the third day postimplantation. Sixteen of 25 juvenile flukes transplanted into chicks' eyes lacking nictitating membranes survived, grew, and developed on the sciera, cornea, and eyelids. Four flukes migrated to intact eyes and developed on the normal site, the interior surface of the nictitating membrane. They were larger than worms that remained in altered eyes. Twenty-three of 120 metacercariae inoculated into the median slits of 7 chicks lacking both nictitating membranes survived, grew, and developed in the altered eyes. Since fertile worms were not recovered from eyes lacking nictitating membranes, the possibility exists that the physicochemical requirements of sexually mature eye flukes differ from those of juvenile flukes.

Comparison of bretylium and guanethidine: tolerance, and effects on adrenergic nerve function and responses to sympathomimetic amines.

Bretylium depresses the slope of regression lines relating frequency of sympathetic nerve stimulation to magnitude of contractions of the cat nictitating membrane. In contrast, guanethidine and reserpine preferentially abolish responses to low rates of nerve stimulation and cause a roughly parallel shift of the regression lines. The hypersensitivity of the nictitating membranes of cats to intravenous adrenaline or noradrenaline is far greater after a series of small daily doses of bretylium or guanethidine than after single large doses. The maximal sensitivity produced was similar to that after postganglionic sympathetic nerve section and exceeded that produced by ganglion blockade. The development of hypersensitivity to catechol amines is accompanied by some return of responses of the nictitating membranes to sympathetic nerve stimulation despite continued daily administration of bretylium or guanethidine. In cats given bretylium daily, responses to low rates of nerve stimulation become greater than in controls unless the dose of bretylium given subcutaneously is 50 mg/kg or more. When marked hypersensitivity to catechol amines has been produced by giving bretylium or guanethidine daily for 7 or 14 days, the sympathomimetic effects of these compounds are greater. Responses to intravenous dimethylphenylpiperazinium are also increased and the results suggest that even large daily doses of adrenergic neurone blocking agents do not appreciably impair the functioning of the adrenal medulla. The pressor effects of intravenous adrenaline, noradrenaline and dimethylphenylpiperazinium iodide increase less than the corresponding nictitating membrane responses. These results are discussed in relation to tolerance to adrenergic neurone blockade, and differences between the effects of bretylium and guanethidine found in man. Bretylium and guanethidine depress the slopes of the dose-response curves for the pressor and nictitating membrane contracting effects of tyramine. When single doses or a short series of daily doses were given, guanethidine caused more depression of the slopes than did bretylium, but nevertheless large depressions of slope were found after giving bretylium daily for several weeks. The magnitude of the responses can be greater or less than in controls depending on the dose of the sympathomimetic amine, the dose of the adrenergic neurone blocking agent and the duration of its administration. The results suggest that injection of tyramine produces a progressively smaller release of adrenaline or noradrenaline during the daily administration of bretylium (or guanethidine) but that in some test situations this is more than compensated for by the development of hypersensitivity to the catechol amine released. Some corresponding changes in responses to amphetamine and ephedrine are also described.

自律神經毒作用知見補遺

Responses of normal and sympathetic denervated nictitating membranes and pupils (in situ), and of the sphincter and dilatator iridis muscles (in vitro) of cat to various autonomic drugs were comparatively studied. The drugs used were adrenaline (Ad.), noradrenaline (NAd.), Acetylcholine (ACh), pilocarpine, atropine, hexamethonium (C6) and TEA. The peripheral actions of C6 and TEA were analyzed by the effects of these drugs upon the action of Ad., NAd. and ACh. From these experimental results the innervation of nictitating membranes and pupils was discussed.

一定藥物の作用と副腎中のAdrenalin及びNoradrenalinの産生について

The investigation on the amounts of adrenaline (A) and noradrenaline (N) in the suprarenal gland and in the blood after administration of mecholyl, acetylcholine, nicotine and the splanchnic stimulation was performed. Rat's blood-pressure and rabbit's intestine were used to estimate the contents of A and N in the isolated rabbit's suprarenal gland, the Gaddum's formula being applied for calculation. The proportions of A and N liberated from the suprarenal gland were determined by the method. of Bülbring and Burn, using a denervated cat's nictitating membranes. Under the conditions mentioned above an increase of N and a decrease of A was observed. From these results it may be considered that N plays an important part as an adrenergic transmitter.

Experimentally-induced petechial hemorrhage and white embolization in the rabbit's nictitating membrane

1. 1. Various hemorrhagic agents were injected intradermally into ear, abdomen, conjunctiva, and nictitating membrane of about three hundred nonanesthetized male rabbits, and biomicroscopic observations were made at the site of injection. 2. 2. Different areas of the skin of the same animal differed in the degree of petechial production with the same test agent. Because of this finding these regions were not considered conducive to any approach of a quantitative nature. The best region found for observational studies of petechial hemorrhage was the nictitating membrane. 3. 3. A new technique which utilizes both nictitating membranes of the rabbit's eyes was developed and is described to test the fragility of the minute blood vessels within a three-hour test period by the time of occurrence and number of petechiae. 4. 4. At a given dosage administered, a correlation was found between the number and the time of occurrence of petechiae. When different dosages were used, the number produced correlated directly with the dosage. Physiologic saline, distilled water, and mineral oil served as controls and did not produce petechiae. 5. 5. It was possible to divide the animals into two groups, those susceptible and those resistant to induced hemorrhage by a given test agent. 6. 6. Individual variations were found in the reparability of the damaged vessels. Some animals repaired these vessels in a few hours, while others required days. 7. 7. The following agents were used locally to produce petechial hemorrhage in the nictitating membrane: Merthiolate, Shiga protein toxin, moccasin snake venom, croton oil in mineral oil, sodium heparin, heparin plasma, saline plasma, citrate plasma, serum, clostridium welchii toxin, clostridium septicum toxin, streptolysin streptococcus pyogenes Group A, cysteine hydrochloride, and tricresol. 8. 8. Capillary ruptures induced by croton oil are probably due to the formation of large particles which embolize. The embolization of these vessels leads to their rupture and petechial manifestation. 9. 9. The local or systemic injection of heparin results in the constant occurence of white emboli in the nictitating membrane. It appeared that these emboli were of a different nature than those produced by croton oil. 10. 10. The local injection of heparin always resulted in petechiae in the nictitating membrane, although the systemic injection of heparin alone did not necessarily result in petechiae. Systemic heparinization combined with Merthiolate always produced petechiae in the nictitating membrane. 11. 11. When local application of Merthiolate or croton oil was preceded by the systemic injection of sodium heparin, more petechial hemorrhages resulted. 12. 12. The first occurrence of white emboli preceded or was observed simultaneously with the first occurrence of petechiae. Such emboli were not, or only rarely, observed following injections of moccasin snake venom and Merthiolate into the nictitating membrane, although innumerable petechiae occurred with these agents. 13. 13. Serum, heparin plasma, and citrate plasma, from rabbit origin, produced white emboli and petechiae following local injection in the nictitating membrane. With the exception of citrate plasma, petechiae were always produced. The latter appeared to diminish petechial formation and also the appearance of white emboli, both of which were lacking in the nictitating membrane of one animal but present in the four membranes of two other animals.