In Vivo Confocal Microscopy of Prominent Conjunctival and Corneal Nerves in Multiple Endocrine Neoplasia Type 2B.

OBJECTIVE: To report a case of a patient affected by multiple endocrine neoplasia type 2B (MEN 2B) with imaging of conjunctival neuromas by in vivo confocal microscopy (IVCM). METHODS: Case report. RESULTS: A 48-year-old patient affected by MEN2B complained of progressive visual loss in his right eye and severe red, dry and itchy eyes. Best-corrected visual acuity was 20/63 OD and 20/25 OS. Slit lamp exam showed thickened and turned out lid margins, significant blepharitis, conjunctival injection, multiple presumed subconjunctival neuromas at the bulbar conjunctiva and at the limbus, marked prominence of corneal nerves, exposure keratopathy due to incomplete blinking and corneal hypoesthesia, subepithelial corneal neovascularization and scarring in the mid inferior part of both corneas and bilateral iris nodules. We performed IVCM on conjunctival neuromas, revealing large, thick bundles of nerves with disorganization, prominent loops, bifurcations and dilations measuring as much as 1 mm. The IVCM of corneal nerves demonstrated hypertrophic sub basal plexus. CONCLUSIONS: To date, this is the first report which documents conjunctival neuromas by confocal microscopy in MEN2B.

Anatomical and functional dichotomy of ocular itch and pain.

Itch and pain are refractory symptoms of many ocular conditions. Ocular itch is generated mainly in the conjunctiva and is absent from the cornea. In contrast, most ocular pain arises from the cornea. However, the underlying mechanisms remain unknown. Using genetic axonal tracing approaches, we discover distinct sensory innervation patterns between the conjunctiva and cornea. Further genetic and functional analyses in rodent models show that a subset of conjunctival-selective sensory fibers marked by MrgprA3 expression, rather than corneal sensory fibers, mediates ocular itch. Importantly, the actions of both histamine and nonhistamine pruritogens converge onto this unique subset of conjunctiva sensory fibers and enable them to play a key role in mediating itch associated with allergic conjunctivitis. This is distinct from skin itch, in which discrete populations of sensory neurons cooperate to carry itch. Finally, we provide proof of concept that selective silencing of conjunctiva itch-sensing fibers by pruritogen-mediated entry of sodium channel blocker QX-314 is a feasible therapeutic strategy to treat ocular itch in mice. Itch-sensing fibers also innervate the human conjunctiva and allow pharmacological silencing using QX-314. Our results cast new light on the neural mechanisms of ocular itch and open a new avenue for developing therapeutic strategies.

In Vivo Small Molecule Delivery to the Optic Nerve in a Rodent Model.

Small molecule delivery to the optic nerve would allow for exploration of molecular and cellular pathways involved in normal physiology and optic neuropathies such as glaucoma, and provide a tool for screening therapeutics in animal models. We report a novel surgical method for small molecule drug delivery to the optic nerve head (ONH) in a rodent model. In proof-of-principle experiments, we delivered cytochalasin D (Cyt D; a filamentous actin inhibitor) to the junction of the superior optic nerve and globe in rats to target the actin-rich astrocytic cytoskeleton of the ONH. Cyt D delivery was quantified by liquid chromatography and mass spectrometry of isolated optic nerve tissue. One day after Cyt D delivery, anterior ONH filamentous actin bundle content was significantly reduced as assessed by fluorescent-tagged phalloidin labeling, relative to sham delivery. Anterior ONH nuclear counts and axon-specific beta-3 tubulin levels, as well as peripapillary retinal ganglion cell layer nuclear counts were not significantly altered after Cyt D delivery relative to sham delivery. Lastly, the surgical delivery technique caused minimal observable axon degeneration up to 10 days post-surgery. This small molecule delivery technique provides a new approach to studying optic neuropathies in in vivo rodent models.

Topography of the supraorbital nerve with reference to the lacrimal caruncle: danger zone for direct browplasty.

PURPOSE: To elucidate the course of the supraorbital nerve (SON) with reference to the lacrimal caruncle in order to facilitate safer direct browplasty by preventing nerve injury. METHODS: Thirty-four hemifaces from 18 embalmed Korean cadavers were dissected. A vertical line through the upmost point of the lacrimal caruncle and a horizontal line through the supraorbital margin were used as the horizontal and vertical reference positions, respectively. The course of the SON in the frontal view and the point at which it pierced the overlaying musculature were examined. RESULTS: The SON divides into a superficial branch and a deep branch just after exiting the orbit. In all cases, the deep SON remains in the subgaleal plane deep to the corrugator and frontalis muscles. The superficial SON travels under the corrugator muscle dividing into three branches (medial, intermediate and lateral) and pierced the frontalis muscle at 19-32 mm above the supraorbital margin. However, in 11 cases (32%) the medial branch of the superficial SON pierced the lower portion of the corrugator muscle at 3.6 mm above the supraorbital margin and ran in front of the muscle along with the vertical line through the upmost point of the lacrimal caruncle. CONCLUSIONS: One-third of the medial branch of the superficial SON without corrugator muscle protection is vulnerable to iatrogenic injury during direct browplasty. Therefore, the oculofacial surgeon must bring the dissection plane of the forehead tissue more superficially around the vertical line through the upmost point of the lacrimal caruncle in order to avoid nerve injury.

The utility of an artificial substitute to improve corneal sensitivity in glaucomatous patients on chronic therapy with prostaglandin analogs.

PURPOSE: To evaluate whether a tear substitute can improve corneal subepithelial nerve plexus and corneal sensitivity in glaucomatous patients. METHODS: This study was prospective, longitudinal, and single arm. Twenty glaucomatous patients were recruited. All the patients were treated with a prostaglandin analog with preservative for at least 1 year. Preservative-free artificial tears (0.5% tamarind seed polysaccharide 0.5(®) eye drops single-dose preservative free [Oftagen]) were prescribed thrice per day. The participants were subjected to clinical and instrumental evaluation at baseline (T0), after 1 month (T1) and after 3 months (T3) of treatment. All patients were examined with a digital corneal confocal laser-scanning microscope (HRT II Rostock Cornea Module; Heidelberg Engineering GmbH) and Cochet-Bonnet corneal esthesiometer. RESULTS: After the artificial substitute, corneal and conjunctival sensitivity significantly (P < 0.001) improved at T1 and T3 compared to the baseline. A significant correlation was found between break-up time and both central corneal sensitivity and the number of fibers. CONCLUSION: The use of a preservative-free artificial substitute in association with a topical therapy with chronic preservative could increase the compliance of patients.

Ocular surface sensitivity repeatability with Cochet-Bonnet esthesiometer.

PURPOSE: To determine the repeatability of ocular surface threshold measurements using the Cochet-Bonnet esthesiometer on the same day and 3 months apart. METHODS: Two separate studies were conducted to determine the repeatability of ocular surface threshold measurements made on the same day (n = 20 subjects) and 3 months apart (n = 29 subjects). The Cochet-Bonnet esthesiometer was used to measure corneal and inferior conjunctival thresholds using the ascending method of limits. The pressure exerted by the Cochet-Bonnet esthesiometer was determined using an analytical balance, for both the 0.08- and 0.12-mm-diameter filaments. This calibration was then used to convert filament length measurements to pressure. Repeatability was determined using a Bland and Altman analysis. RESULTS: The pressure exerted at each filament length differed between the two filament diameters. The measured pressure also differed from values provided by the manufacturer. Repeatability of threshold measurements at the central cornea was shown to be good, with better repeatability for same-day measurements (coefficient of repeatability [CoR] = ±0.23 g/mm²) than for those 3 months apart (CoR = ±0.52 g/mm²). Threshold measurements at the inferior conjunctiva, in contrast, were poorly repeatable (CoR = ±12.78 g/mm²). CONCLUSIONS: Cochet-Bonnet esthesiometry is repeatable when performed on the central cornea on the same day and 3 months apart, but this instrument is not recommended for conjunctival threshold measurements.

Bilateral nerve alterations in a unilateral experimental neurotrophic keratopathy model: a lateral conjunctival approach for trigeminal axotomy.

To study bilateral nerve changes in a newly developed novel mouse model for neurotrophic keratopathy by approaching the trigeminal nerve from the lateral fornix. Surgical axotomy of the ciliary nerve of the trigeminal nerve was performed in adult BALB/c mice at the posterior sclera. Axotomized, contralateral, and sham-treated corneas were excised on post-operative days 1, 3, 5, 7 and 14 and immunofluorescence histochemistry was performed with anti-ß-tubulin antibody to evaluate corneal nerve density. Blink reflex was evaluated using a nylon thread. The survival rate was 100% with minimal bleeding during axotomy and a surgical time of 8±0.5 minutes. The blink reflex was diminished at day 1 after axotomy, but remained intact in the contralateral eyes in all mice. The central and peripheral subbasal nerves were not detectable in the axotomized cornea at day 1 (p<0.001), compared to normal eyes (101.3±14.8 and 69.7±12.0 mm/mm² centrally and peripherally). Interestingly, the subbasal nerve density in the contralateral non-surgical eyes also decreased significantly to 62.4±2.8 mm/mm² in the center from day 1 (p<0.001), but did not change in the periphery (77.3±11.7 mm/mm², P = 0.819). Our novel trigeminal axotomy mouse model is highly effective, less invasive, rapid, and has a high survival rate, demonstrating immediate loss of subbasal nerves in axotomized eyes and decreased subbasal nerves in contralateral eyes after unilateral axotomy. This model will allow investigating the effects of corneal nerve damage and serves as a new model for neurotrophic keratopathy.

Effect of VIP on intracellular [Ca2+], extracellular regulated kinase 1/2, and secretion in cultured rat conjunctival goblet cells.

PURPOSE: To determine the intracellular signaling pathways that vasoactive intestinal peptide (VIP) uses to stimulate high molecular weight glycoconjugate secretion from cultured rat conjunctival goblet cells. METHODS: Goblet cells from rat bulbar and forniceal conjunctiva were grown in organ culture. Presence and localization of VIP receptors (VPAC1 and 2) were determined by RT-PCR, immunofluorescence microscopy and Western blot analysis. Intracellular [Ca(2+)] ([Ca(2+)]i) was measured using fura-2. Extracellular signal-regulated kinase (ERK)-1/2 activity was determined by Western blot analysis. High molecular weight glycoconjugate secretion was measured with an enzyme-linked lectin assay on cultured goblet cells that were serum-starved for 2 hours before stimulation with VIP, VPAC1-, or VPAC2-specific agonists. Inhibitors were added 30 minutes prior to VIP. Activation of epidermal growth factor receptor (EGFR) was measured by immunoprecipitation using an antibody against pTyr followed by Western blot analysis with an antibody against EGFR. RESULTS: Both VIP receptors were present in rat conjunctiva and cultured goblet cells. VIP- and VPAC-specific agonists increased [Ca(2+)]i and secretion in a concentration-dependent manner. VIP also increased ERK1/2 activity, VIP-stimulated increase in [Ca(2+)]i. Secretion, but not ERK1/2 activity, was inhibited by the protein kinase A inhibitor, H89. VIP-stimulated secretion was inhibited by siRNA for ERK2 but not by siRNA for EGFR. VIP did not increase the phosphorylation of the EGFR. CONCLUSIONS: In conclusion, in cultured rat conjunctival goblet cells, VPAC1 and 2 receptors are functional. VIP stimulates a cAMP-dependent increase in [Ca(2+)]i and glycoconjugate secretion, but not ERK1/2 activation. VIP does not activate with EGFR.

Corneal and conjunctival sensory function: the impact on ocular surface sensitivity of change from low to high oxygen transmissibility contact lenses.

PURPOSE: Deprivation of oxygen to the ocular surface during contact lens wear has been implicated in the alteration of sensory function. This study investigates whether increasing oxygen availability through discontinuation of contact lens wear or transfer into highly oxygen transmissible (high Dk/t) lenses leads to a change in corneal or conjunctival sensitivity. METHODS: Twenty-seven long-term extended wearers of low Dk/t soft contact lenses ceased lens wear for 1 week and were refitted with high Dk/t silicone hydrogel lenses. A control group of 25 nonwearers matched for age and sex was also recruited. Central corneal and inferior conjunctival sensitivity were measured using an air-jet aesthesiometer. Threshold was determined using a staircase technique. Measurements were taken during low Dk/t lens wear; after 1 week of no wear; and after 1, 3, 6, and 12 months of high Dk/t lens wear. Measurements were carried out on one occasion on the nonwearers. RESULTS: Corneal sensitivity decreased 1 week after discontinuation of low Dk/t lenses and no further change in sensitivity occurred with high Dk/t lens wear. Conjunctival sensitivity did not change over the same time frame. Ocular surface sensitivity in long-term low Dk/t soft lens wearers was similar to that of nonwearers. Sensitivity was higher in females than males in the nonwearers, but not in the lens-wearing group. An interaction of sex on change in conjunctival threshold was found in the lens wearers. CONCLUSIONS: These findings indicate that factors other than oxygen availability alone determine sensitivity of the ocular surface. Silicone hydrogel contact lenses appear to have only a minor impact on ocular surface sensitivity in previous lens wearers.

New "drug-in cyclodextrin-in deformable liposomes" formulations to improve the therapeutic efficacy of local anaesthetics.

The combined approach of cyclodextrin complexation and entrapment in liposomes was investigated to develop a topical formulation of local anaesthetics. For both benzocaine (BZC) and butamben (BTM), hydroxypropyl-beta-cyclodextrin (HPbetaCD) was a better partner than betaCD; drug-HPbetaCD coevaporated products showed the best solubility and dissolution properties, and were selected for loading into liposomes. Addition of stearylamine to the phosphatidylcholine-cholesterol mixture of the vesicle bilayer allowed obtainment of deformable liposomes with improved permeation and in vivo drug anaesthetic effect (P<0.05). Double-loaded deformable liposomes were obtained by adding the drug-HPbetaCD complex at its maximum aqueous solubility in the vesicles hydrophilic phase, and the remaining amount up to 1% as free drug in the lipophilic phase. The properties of double-loaded liposomes were compared with those of classic single-loaded ones, obtained by adding 1% free drug in the aqueous or lipophilic phase of the vesicles. Size, charge, morphology and entrapment efficiency of the different batches were investigated, respectively, by light scattering, confocal laser scanning microscopy and dialysis, while their therapeutic efficacy was evaluated in vivo on rabbits. For both drugs, double-loaded liposomes, exploiting the favourable effects of drug-CD complexation, allowed a significant (P<0.05) enhancement of intensity and duration of anaesthetic effect with respect to those single-loaded.

Distinct central representations for sensory fibers innervating either the conjunctiva or cornea of the rat.

The laminar sheet of epithelium (e.g., skin and mucous membrane) enclosing our bodies is represented in the dorsal horns of the medulla and spinal cord. The eyeball however indents this laminar sheet and is shrouded by different layers: the cornea/sclera, the conjunctiva, and hairy skin. This involution of the orb confounds defining the central representation of the cornea and its surrounding mucosa and skin. We used herein the transganglionic transport of a cocktail of HRP conjugated to cholera toxin and wheat germ agglutinin to determine the central representation of these epithelia in the dorsal horns of the rat. The HRP cocktail was injected either into the stroma of the cornea, the mucosa of the conjunctiva, or the supraorbital and infraorbital nerves. Injections of the cornea produced dense label in the interstitial islands in the ventral medullary dorsal horn (MDH), probably lamina I, and in neuropil in the ventromedial tip of the MDH, probably lamina II. There sometimes was variable, diffuse label in the C1 dorsal horn after corneal injections but more rostral parts of the trigeminal sensory complex were never labeled. Injections of the conjunctiva densely labeled laminae I-III in the C1 dorsal horn, while laminae IV-V were diffusely labeled. Sparser reaction product also was seen in lamina I in positions similar to the cornea projection. Label was seen ventrally in subnuclei interpolaris and oralis, as well as the principal trigeminal nucleus. Projections of the infraorbital nerve included all laminae in the trigeminocervical complex as well as large portions of the rostral subnuclei in the spinal trigeminal nucleus. The projections of the supraorbital nerve were similar, but were restricted to ventral parts of the trigeminal sensory complex. In other cases the cornea was injected either after cutting the supraorbital and infraorbital nerves or the conjunctiva was injected after enucleating the eyeball. Any reaction product from corneal injections was reduced dramatically in the C1 dorsal horn after transection of the infraorbital and supraorbital nerves. Injecting the conjunctiva after enucleating the eyeball densely labeled the C1 projection to the dorsal horn, a small patch in lamina I in the MDH, as well as the rostral trigeminal complex. We propose that the cornea has but a single representation in the trigeminocervical complex in its ventral part near the caudal end of the medulla. We also propose the palpebral conjunctiva mucosa is represented in the C1 dorsal horn, and speculate that the bulbar conjunctiva overlaps with that of the cornea in lamina I. We discuss these projections in relation to the circuitry for the supraorbital-evoked and corneal-evoked blink reflexes. The relationship of the cornea and conjunctiva is intimate, and investigators must be very careful when attempting to stimulate them in isolation.

Frequent spontaneous eyeblink activity associated with reduced conjunctival surface (trigeminal nerve) tactile sensitivity.

BACKGROUND: A number of recent studies have reported high spontaneous eyeblink rate (SEBR) values in apparently normal subjects, but the reasons for this are unclear. METHODS: An assessment was made of SEBR, in 60 educated adult male subjects aged between 22 and 40 years, over a period of 5 min in silence. Half of the subjects were classified as having frequent eyeblink activity. All subjects also had their corneal and conjunctival touch (tactile) sensitivity assessed with a Cochet-Bonnet aesthesiometer immediately after the video recording of SEBR. RESULTS: SEBR ranged from 4.6 to 43.5 (mean 18.6) eyeblinks/min. The SEBR was 26.8 +/- 6.0 eyeblinks/min for those with frequent eyeblink activity as compared to just 10.3 +/- 3.5 eyeblinks/min for those with normal eyeblink activity (p < 0.001). There was no difference in palpebral aperture or exposed ocular surface area between the two groups. The average central corneal sensitivity was only marginally different between the two groups (56.8 +/- 2.8 mm vs 58.5 +/- 2.3 mm) but the conjunctival threshold sensitivity was substantially different (at 23.8 +/- 4.3 mm vs 28.5 +/- 3.5 mm; p < 0.001). SEBR was inversely correlated with the conjunctival sensitivity in those with frequent eyeblink activity (p < 0.001). CONCLUSIONS: Our study provides a clue as to the mechanism of inhibition of spontaneous eyeblink activity, namely that a certain level of ocular surface (conjunctival) sensitivity is required to keep SEBR low.

Responsiveness of short-lasting unilateral neuralgiform headache with conjunctival injection and tearing to hypothalamic deep brain stimulation.

Short-lasting unilateral neuralgiform headache with conjunctival injection and tearing (SUNCT) is a severe primary headache disorder that is often refractory to medical therapy. Although the pathogenesis of this and other trigeminal autonomic cephalalgias is not completely understood, ipsilateral activation of the posterior and inferior hypothalamus has been identified on functional imaging studies during attacks. The authors report on a case of SUNCT syndrome successfully treated with hypothalamic deep brain stimulation and discuss the current literature.

Nerve growth factor eye drop administrated on the ocular surface of rodents affects the nucleus basalis and septum: biochemical and structural evidence.

It has been shown that conjunctivally applied NGF in rats can reach the retina and optic nerve. Whether topical eye NGF application reaches the central nervous system is not known. In the present study, we have addressed this question. It was found that topical eye NGF application affects brain cells. Time-course studies revealed that repeated NGF application leads to high concentration of this neurotrophins after 6 h and normal levels after 24 h. Our results also showed that topical eye application of NGF causes an enhanced expression of NGF receptors and ChAT immunoreactivity in forebrain cholinergic neurons, suggesting that ocular NGF application could have a functional role on damaged brain cells. The present findings suggest that eye NGF application can represent an alternative route to prevent degeneration of NGF-receptive neurons involved in disorders such as Alzheimer and Parkinson.

Autonomic features in cluster headache. Exploratory factor analysis.

The objective is to identify the pathogenesis of each autonomic manifestation in cluster headache (CH). Through a deductive statistics method (factor analysis) we analysed the type of autonomic symptoms reported by 157 CH patients. Three principal components were identified in the analysis: parasympathetic activation (lacrimation, conjunctival injection and rhinorrhoea), sympathetic defect (miosis and ptosis) and parasympathetic mediated effect (nasal congestion, eyelid oedema and forehead sweating). This work suggests that there are three different mechanisms underlying autonomic manifestations in CH.

The inhibitory interaction between human corneal and conjunctival sensory channels.

PURPOSE: To explore human corneal and conjunctival sensory channels at suprathreshold level. METHODS: Ten healthy human subjects participated in the study. The Belmonte pneumatic esthesiometer was used to apply mechanical and chemical stimuli to the central cornea and temporal conjunctiva of the left eye. Stimuli were applied in a paired and unpaired way for conjunctival stimulation. A 100-point visual analog scale (VAS) was used to rate the intensity of the stimulus. RESULTS: The magnitudes of the sensation evoked from the conjunctiva were different when using different methods for presenting stimuli to the ocular surface. When stimuli were applied to the conjunctiva alone, the magnitude of the sensation was stronger than when the stimuli were applied in pairs to the cornea and conjunctiva for both mechanical (P = 0.04) and chemical (P = 0.02) stimulation. CONCLUSIONS: The relatively strong discomfort evoked from the cornea appears to suppress partially the relatively weaker conjunctival stimulation. This manifested as the conjunctival sensory transducer function being shallower (less intense sensation) when immediately preceded by corneal stimulation than when the conjunctival sensory transducer functions were measured alone (unpaired). The underlying mechanism could be adaptation or some other inhibitory effect, such as diffuse noxious inhibitory control. At some level therefore, corneal and conjunctival sensory channels are not independent.

Substance P and opioid peptidergic innervation of the anterior eye segment of the rat: an immunohistochemical study.

Recently discovered endogenous opioid peptides such as nociceptin are known to modulate neurotransmitter release of primary afferent neurons (especially substance P, SP) and they have also been demonstrated in peripheral nerve fibres. The aim of this study was to investigate the opioid peptidergic innervation of the anterior eye segment and to compare it with the innervation pattern of SP in order to shed light on the functional relationship between these peptides. Anterior eye segments of 20 rat eyes were cut in a tangential plane and the sections stained with antibodies against SP, nociceptin, nocistatin, endomorphin 1 and 2, leu-enkephalin and met-enkephalin. Sections of the spinal cord or brain were used as positive controls. Numerous SP-immunoreactive nerve fibres were found in the conjunctiva, cornea, episclera, trabecular meshwork, iris and ciliary body. A weak staining for met-enkephalin and leu-enkephalin could only be found in the iris and anteriormost ciliary body. Nerve fibres immunoreactive for nociceptin, nocistatin, and endomorphin 1 or 2 could not be detected in any part of the anterior eye segment. It is tempting to speculate that the opioid peptidergic innervation of the anterior ciliary body may play a role in the modulation of intraocular inflammation.

Cellular inflammatory response induced by sensory denervation of the conjunctiva in monkeys.

The inflammatory response induced by sensory denervation of the cornea, neuroparalytic (neurotrophic) keratitis, has been widely reported in the literature. Clinical evidence has shown that the conjunctiva also responds to sensory denervation, but little is known of the cytology of the conjunctival tissue response to denervation. The purpose of this study therefore was to investigate the cytological aspects of tissue response induced in the conjunctiva of monkeys by sensory denervation. Intracranial ophthalmic neurotomy was carried out in three monkeys, maxillary neurotomy in four, combined ophthalmic and maxillary neurotomy in two, and infraorbital nerve transaction in one monkey. These various operations were performed for other experimental purposes, but the conjunctival tissues from the animals were suitable and available to study the cytology of the inflammatory response induced in the tissue following the sensory denervation. The cytological changes were studied using light microscopy. Complete or severe ophthalmic nerve transection induced significant inflammatory responses, which were largely confined to the tarsal region of the conjunctiva. The responses included a substantial increase in the infiltration of the epithelium by polymorphonuclear leucocytes and severe disruption of the epithelium. The number of macrophages in the conjunctiva also increased. The response induced by maxillary denervation, however, was not as pronounced as that induced by sensory denervation. The restriction of the conjunctival tissue response to the tarsal region was considered to be due to the friction between the tarsal conjunctiva (and the hard tarsal plate) and the cornea during blinking. This study shows that sensory denervation of the conjunctiva elicits an inflammatory response characterized by substantial infiltration of the epithelium by neutrophil and macrophage and disorganization of the conjunctival tissue.

[Changes of the conjunctival nervous fillets in diabetic patients]. / Modificari ale filetelor nervoase in conjunctiva bolnavilor cu diabet zaharat.

PURPOSE: Our study suggested to reveal nervous fillets changes from conjunctiva structure in patients with diabetes. MATERIAL AND METHOD: We removed bulbar conjunctiva during cataract surgery on diabetic and non-diabetic patients from the same age group. The conjunctiva fragments were fixed in Lillie's solution, then studied by using optical microscopy with usual and histochemical stainings. RESULTS: Nervous fillets just like perinervium are better represented in younger diabetic patients without diabetic retinopathy or with early diabetic retinopathy and non-diabetic patients, too. In older patients and old diabetes, nervous fillets are much less in number with poor represented perinervium. CONCLUSIONS: Significant changes of the conjunctiva nervous fillets were found in patients with long-term diabetes.

Tear secretion induced by selective stimulation of corneal and conjunctival sensory nerve fibers.

PURPOSE: To measure the increase in tear secretion evoked by selective stimulation of the different populations of sensory receptors of the cornea and conjunctiva by using moderate and intense mechanical, chemical, and cold stimuli. METHODS: Six healthy subjects participated in the study. Tear secretion was measured in both eyes by the Schirmer's test conducted under control conditions and after stimulation of the center of the cornea and the temporal conjunctiva with a gas esthesiometer. Mechanical stimulation consisted in three pulses of 3 seconds' duration of warmed air (at 34 degrees C on the eye surface) applied at moderate (170 mL/min) and high (260 mL/min) flow rates. Cold thermal stimulation was made with cooled air that produced a corneal temperature drop of -1 degrees C or -4.5 degrees C. Chemical (acidic) stimulation was performed with a jet of gas containing a mixture of 80% CO(2) in air. RESULTS: The basal volume of tear secretion increased significantly (P < 0.05, paired t-test) after stimulation of the cornea with high-flow mechanical stimuli (260 mL/min), intense cooling pulses (-4.5 degrees C), and chemical stimulation (80% CO(2)). The same stimuli were ineffective when applied to the conjunctiva. Moderate mechanical (170 mL/min) and cold (-1 degrees C) stimulation of the cornea or the conjunctiva did not change significantly the volume of tear secretion. CONCLUSIONS: Reflex tear secretion caused by corneal stimulation seems to be chiefly due to activation of corneal polymodal nociceptors, whereas selective excitation of corneal mechanonociceptors or cold receptors appears to be less effective in evoking an augmented lacrimal secretion. Conjunctival receptors stimulated at equivalent levels do not evoke an increased tear secretion.