Publications by authors named "Juana Gallar"

47 Publications

Unilateral Corneal Insult Also Alters Sensory Nerve Activity in the Contralateral Eye.

Front Med (Lausanne) 2021 15;8:767967. Epub 2021 Nov 15.

Instituto de Neurociencias, Universidad Miguel Hernández-CSIC, San Juan de Alicante, Spain.

After the unilateral inflammation or nerve lesion of the ocular surface, the ipsilateral corneal sensory nerve activity is activated and sensitized, evoking ocular discomfort, irritation, and pain referred to the affected eye. Nonetheless, some patients with unilateral ocular inflammation, infection, or surgery also reported discomfort and pain in the contralateral eye. We explored the possibility that such altered sensations in the non-affected eye are due to the changes in their corneal sensory nerve activity in the contralateral, not directly affected eye. To test that hypothesis, we recorded the impulse activity of the corneal mechano- and polymodal nociceptor and cold thermoreceptor nerve terminals in both eyes of guinea pigs, subjected unilaterally to three different experimental conditions (UV-induced photokeratitis, microkeratome corneal surgery, and chronic tear deficiency caused by removal of the main lacrimal gland), and in eyes of naïve animals . Overall, after unilateral eye damage, the corneal sensory nerve activity appeared to be also altered in the contralateral eye. Compared with the naïve guinea pigs, animals with unilateral UV-induced mild corneal inflammation, showed on both eyes an inhibition of the spontaneous and stimulus-evoked activity of cold thermoreceptors, and increased activity in nociceptors affecting both the ipsilateral and the contralateral eye. Unilateral microkeratome surgery affected the activity of nociceptors mostly, inducing sensitization in both eyes. The removal of the main lacrimal gland reduced tear volume and increased the cold thermoreceptor activity in both eyes. This is the first direct demonstration that unilateral corneal nerve lesion, especially ocular surface inflammation, functionally affects the activity of the different types of corneal sensory nerves in both the ipsilateral and contralateral eyes. The mechanisms underlying the contralateral affectation of sensory nerves remain to be determined, although available data support the involvement of neuroimmune interactions. The parallel alteration of nerve activity in contralateral eyes has two main implications: a) in the experimental design of both preclinical and clinical studies, where the contralateral eyes cannot be considered as a control; and, b) in the clinical practice, where clinicians must consider the convenience of treating both eyes of patients with unilateral ocular conditions to avoid pain and secondary undesirable effects in the fellow eye.
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http://dx.doi.org/10.3389/fmed.2021.767967DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8634144PMC
November 2021

Small fiber neuropathy in the cornea of Covid-19 patients associated with the generation of ocular surface disease.

Ocul Surf 2021 Nov 12;23:40-48. Epub 2021 Nov 12.

Instituto Oftalmológico Fernández-Vega, Oviedo, Spain; Instituto Universitario Fernández-Vega, Universidad de Oviedo & Fundación de Investigación Oftalmológica, Oviedo, Spain; Instituto de Investigación Sanitaria del Principado de Asturias (ISPA), Oviedo, Spain; Department of Surgery and Medical-Surgical Specialties, Universidad de Oviedo, Oviedo, Spain.

Purpose: To describe the association between Sars-CoV-2 infection and small fiber neuropathy in the cornea identified by in vivo corneal confocal microscopy.

Methods: Twenty-three patients who had overcome COVID-19 were recruited to this observational retrospective study. Forty-six uninfected volunteers were also recruited and studied as a control group. All subjects were examined under in vivo confocal microscopy to obtain images of corneal subbasal nerve fibers in order to study the presence of neuroma-like structures, axonal beadings and dendritic cells. The Ocular Surface Disease Index (OSDI) questionnaire and Schirmer tear test were used as indicators of Dry Eye Disease (DED) and ocular surface pathology.

Results: Twenty-one patients (91.31%) presented alterations of the corneal subbasal plexus and corneal tissue consistent with small fiber neuropathy. Images from healthy subjects did not indicate significant nerve fiber or corneal tissue damage. Eight patients reported increased sensations of ocular dryness after COVID-19 infection and had positive DED indicators. Beaded axons were found in 82.60% of cases, mainly in patients reporting ocular irritation symptoms. Neuroma-like images were found in 65.22% patients, more frequently in those with OSDI scores >13. Dendritic cells were found in 69.56% of patients and were more frequent in younger asymptomatic patients. The presence of morphological alterations in patients up to 10 months after recovering from Sars-CoV-2 infection points to the chronic nature of the neuropathy.

Conclusions: Sars-CoV-2 infection may be inducing small fiber neuropathy in the ocular surface, sharing symptomatology and morphological landmarks with DED and diabetic neuropathy.
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http://dx.doi.org/10.1016/j.jtos.2021.10.010DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8588585PMC
November 2021

Deciphering the Action of Perfluorohexyloctane Eye Drops to Reduce Ocular Discomfort and Pain.

Front Med (Lausanne) 2021 26;8:709712. Epub 2021 Oct 26.

Cellular and Systems Neurobiology Unit, Instituto de Neurociencias, Universidad Miguel Hernández-Consejo Superior de Investigaciones Científicas, San Juan de Alicante, Spain.

Perfluorohexyloctane (F6H8) eyedrops have been recently introduced in Europe as a product to treat dry eye disease, based on its ability to reduce tear film instability in Meibomian gland dysfunction and evaporative dry eye disease, although its mechanism of action is still unknown. In the present pilot study, we evaluated the effects of the ocular instillation of a single drop of commercial F6H8 eyedrops in 20 healthy humans (9 women/11 men), measuring: (a) Corneal surface temperature (CST) from infrared video images; (b) tear volume using phenol red threads; (c) blinking frequency; and (d) ocular surface sensations (cold, dryness, pricking, foreign body, burning, itching, gritty, eye fatigue, watering eyes, and light-evoked discomfort sensations; scored using 10 cm Visual Analog Scales), before and 5-60 min after F6H8 or saline treatment. CST decreased and tearing and blinking frequency increased significantly after F6H8 but not after saline solution. When applied unilaterally, CST decreased only in the F6H8-treated eye. No sensations were evoked after F6H8 or saline. The corneal surface temperature reduction produced by topical F6H8 does not evoke conscious ocular sensations but is sufficient to increase the activity of corneal cold thermoreceptors, leading to an increased reflex lacrimation and blinking that may relieve dry eye condition thus reducing ocular discomfort and pain.
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http://dx.doi.org/10.3389/fmed.2021.709712DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8577568PMC
October 2021

Acute Increase in Blood αCGRP at Maximal Exercise and Its Association to Cardiorespiratory Fitness, Carbohydrate Oxidation and Work Performed: An Exploratory Study in Young Men.

Biology (Basel) 2021 Aug 17;10(8). Epub 2021 Aug 17.

Department of Sports Sciences, Centro de Investigación del Deporte, UMH, Alicante Institute for Health and Biomedical Research (ISABIAL), 03202 Elche, Spain.

This study aimed to explore if the acute variations in plasma concentration of α-calcitonin gene-related peptide (αCGRP) induced by a single maximal exercise bout may be associated to cardiorespiratory fitness and carbohydrate oxidation in humans. Twelve young adult Caucasian men (24.3 ± 0.9 years-old; 179.2 ± 1.9 cm of height; 23.9 ± 0.6 kg·m body mass index) performed a graded exercise test. A venous catheter was placed before testing, and blood samples were taken at baseline, maximal effort and recovery. αCGRP was measured in plasma using a commercial double-sandwich enzyme-linked-immunoassay. A two-way repeated measurements ANOVA was used to compare the values obtained at baseline, maximal effort and recovery. In the whole sample, αCGRP increased at maximal effort and its concentration correlated directly, albeit non-significantly, with the muscle mass normalised VO, VCO, carbohydrate oxidation and relative power. Two thirds of the participants showed an increase in αCGRP concentration at maximal effort. Post hoc analysis showed that in these individuals, the muscle mass normalised VO, VCO, carbohydrate oxidation rate and relative power were higher than in the participants lacking this molecular response. Therefore, our data suggest that (a) a majority of young men respond to exercise with an increase in blood αCGRP concentration; and (b) individuals exhibiting this response also show a higher cardiorespiratory fitness, carbohydrate oxidation and work performed. These findings suggest that this neuropeptide could act as an exerkine with potential effects on physical performance.
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http://dx.doi.org/10.3390/biology10080783DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8389686PMC
August 2021

Membrane potential instabilities in sensory neurons: mechanisms and pathophysiological relevance.

Pain 2022 Jan;163(1):64-74

Laboratory of Ion Channel Research, Department of Cellular and Molecular Medicine, KU Leuven, VIB Center for Brain & Disease Research, Leuven, Belgium.

Abstract: Peripheral sensory neurons transduce physicochemical stimuli affecting somatic tissues into the firing of action potentials that are conveyed to the central nervous system. This results in conscious perception, adaptation, and survival, but alterations of the firing patterns can result in pain and hypersensitivity conditions. Thus, understanding the molecular mechanisms underlying action potential firing in peripheral sensory neurons is essential in sensory biology and pathophysiology. Over the past 30 years, it has been consistently reported that these cells can display membrane potential instabilities (MPIs), in the form of subthreshold membrane potential oscillations or depolarizing spontaneous fluctuations. However, research on this subject remains sparse, without a clear conductive thread to be followed. To address this, we here provide a synthesis of the description, molecular bases, mathematical models, physiological roles, and pathophysiological implications of MPIs in peripheral sensory neurons. Membrane potential instabilities have been reported in trigeminal, dorsal root, and Mes-V ganglia, where they are believed to support repetitive firing. They are proposed to have roles also in intercellular communication, ectopic firing, and responses to tonic and slow natural stimuli. We highlight how MPIs are of great interest for the study of sensory transduction physiology and how they may represent therapeutic targets for many pathological conditions, such as acute and chronic pain, itch, and altered sensory perceptions. We identify future research directions, including the elucidation of the underlying molecular determinants and modulation mechanisms, their relation to the encoding of natural stimuli and their implication in pain and hypersensitivity conditions.
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http://dx.doi.org/10.1097/j.pain.0000000000002289DOI Listing
January 2022

Optical Assessment of Nociceptive TRP Channel Function at the Peripheral Nerve Terminal.

Int J Mol Sci 2021 Jan 6;22(2). Epub 2021 Jan 6.

Department of Medical Neurobiology, Institute for Medical Research Israel-Canada, Hadassah School of Medicine, The Hebrew University, Jerusalem 91123, Israel.

Free nerve endings are key structures in sensory transduction of noxious stimuli. In spite of this, little is known about their functional organization. Transient receptor potential (TRP) channels have emerged as key molecular identities in the sensory transduction of pain-producing stimuli, yet the vast majority of our knowledge about sensory TRP channel function is limited to data obtained from in vitro models which do not necessarily reflect physiological conditions. In recent years, the development of novel optical methods such as genetically encoded calcium indicators and photo-modulation of ion channel activity by pharmacological tools has provided an invaluable opportunity to directly assess nociceptive TRP channel function at the nerve terminal.
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http://dx.doi.org/10.3390/ijms22020481DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7825137PMC
January 2021

Sodium Channel Blockers Modulate Abnormal Activity of Regenerating Nociceptive Corneal Nerves After Surgical Lesion.

Invest Ophthalmol Vis Sci 2021 01;62(1)

Instituto de Neurociencias, Universidad Miguel Hernández - CSIC, San Juan de Alicante, Spain.

Purpose: To test the effect of different sodium channel blockers on the electrical activity of corneal nociceptors in intact and surgically injured corneas.

Methods: In anesthetized guinea pigs, a 4-mm diameter corneal flap was performed in one eye at a midstromal depth using a custom-made microkeratome. At different times after surgery (3 hours to 15 days), the electrical activity of corneal nociceptor fibers was recorded from ciliary nerve filaments in the superfused eye in vitro. Mechanical threshold was measured using calibrated von Frey hairs; chemical stimulation was performed applying 30-second CO2 gas pulses. The characteristics of the spontaneous and stimulus-evoked activity of corneal nociceptors recorded from intact and lesioned corneas, before and after treatment with the sodium channel blockers lidocaine, carbamazepine, and amitriptyline, were compared.

Results: No spontaneous or stimulus-evoked impulse activity was detected inside the flap at any of the studied time points. However, both were recorded from mechanonociceptor and polymodal nociceptors fibers in the surrounding corneal tissue, being significantly higher (sensitization) 24 to 48 hours after surgery. In these fibers, none of the tested drugs affected mechanical threshold, but they significantly reduced the CO2 response of polymodal nociceptors of intact and injured corneas. Likewise, they diminished significantly the transient increase in spontaneous and stimulus-evoked activity of sensitized polymodal nociceptors.

Conclusions: Na+ channel blockers decrease the excitability of intact and sensitized corneal nociceptor fibers, thus acting as potential tools to attenuate their abnormal activity, which underlies the spontaneous pain, hyperalgesia, and allodynia often accompanying surgical corneal lesions, as occurs after photorefractive surgery.
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http://dx.doi.org/10.1167/iovs.62.1.2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7797933PMC
January 2021

Topical treatment with a mu opioid receptor agonist alleviates corneal allodynia and corneal nerve sensitization in mice.

Biomed Pharmacother 2020 Dec 6;132:110794. Epub 2020 Oct 6.

Sorbonne Université, INSERM, CNRS, Institut de la Vision, 17 Rue Moreau, F-75012, Paris, France. Electronic address:

Corneal pain is considered to be a core symptom of ocular surface disruption and inflammation. The management of this debilitating condition is still a therapeutic challenge. Recent evidence supports a role of the opioid system in the management of corneal nociception. However, the functional involvement of the mu opioid receptor (MOR) underlying this analgesic effect is not known. We first investigated the expression of the MOR in corneal nerve fibers and trigeminal ganglion (TG) neurons in control mice and a mouse model of corneal inflammatory pain. We then evaluated the anti-nociceptive and electrophysiological effects of DAMGO ([D-Ala,N-Me-Phe,Gly-ol] enkephalin), a MOR-selective ligand. MOR immunoreactivity was detected in corneal nerve fibers and primary afferent neurons of the ophthalmic branch of the TG of naive mice. MOR expression was significantly higher in both structures under conditions of inflammatory corneal pain. Topical ocular administration of DAMGO strongly reduced both the mechanical (von Frey) and chemical (capsaicin) corneal hypersensitivity associated with inflammatory ocular pain. Repeated instillations of DAMGO also markedly reversed the elevated spontaneous activity of the ciliary nerve and responsiveness of corneal polymodal nociceptors that were observed in mice with corneal pain. Finally, these DAMGO-induced behavioral and electrophysiological responses were totally blunted by the topical application of naloxone methiodide, an opioid receptor antagonist. Overall, these results provide evidence that topical pharmacological MOR activation may constitute a therapeutic target for the treatment of corneal pain and improve corneal nerve function to alleviate chronic pain.
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http://dx.doi.org/10.1016/j.biopha.2020.110794DOI Listing
December 2020

Effects of corneal injury on ciliary nerve fibre activity and corneal nociception in mice: A behavioural and electrophysiological study.

Eur J Pain 2019 03 16;23(3):589-602. Epub 2018 Nov 16.

INSERM, CNRS, Institut de la Vision, Sorbonne Université, Paris, France.

Background: Ocular surface diseases are among the most frequent ocular pathologies. Ocular pain following corneal injury is frequently observed in clinic. Corneal sensory innervation is supplied by ciliary nerves derived from ophthalmic division of the trigeminal ganglion.

Methods & Results: Extracellular activity of the mouse ciliary nerve was first used to investigate the corneal responsiveness to chemical, mechanical and thermal stimulations in order to specifically study the responses of polymodal nociceptors, mechano-nociceptors and cold thermoreceptor in a control cornea. Then, in two models of corneal injury (repeated instillations of 0.02% benzalkonium chloride and corneal scraping), we first measured the corneal sensitivity to chemical (eye-wiping test) and mechanical (von Frey filaments) stimulation. Thereafter, we evaluated whether these corneal injuries modified the spontaneous and chemical stimulation-evoked activity of the ciliary nerve. Both models of injury induced a significant corneal chemical hypersensitivity correlated with an increase of the spontaneous activity of the ciliary nerve and a faster response of the ciliary nerve after a chemical stimulation.

Conclusions: Overall, this study provides new insights into the functional aspects of corneal nerve fibre activity in mice after corneal injury. The increase in ciliary nerve activity may thus contribute to the development of ocular pain after corneal damage.

Significance: This study highlights the parallel increase in ciliary nerve activity and corneal sensitivity after corneal injury in mice. The strategy of combining ex vivo electrophysiological recordings of the ciliary nerve in mice and corneal sensitivity measurements therefore helps to uncover the functional aspects of corneal pain.
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http://dx.doi.org/10.1002/ejp.1332DOI Listing
March 2019

Melanopsin expression in the cornea.

Vis Neurosci 2018 01;35:E004

Department of Ophthalmology,School of Medicine,University of California San Francisco,San Francisco,California.

A unique class of intrinsically photosensitive retinal ganglion cells in mammalian retinae has been recently discovered and characterized. These neurons can generate visual signals in the absence of inputs from rods and cones, the conventional photoreceptors in the visual system. These light sensitive ganglion cells (mRGCs) express the non-rod, non-cone photopigment melanopsin and play well documented roles in modulating pupil responses to light, photoentrainment of circadian rhythms, mood, sleep and other adaptive light functions. While most research efforts in mammals have focused on mRGCs in retina, recent studies reveal that melanopsin is expressed in non-retinal tissues. For example, light-evoked melanopsin activation in extra retinal tissue regulates pupil constriction in the iris and vasodilation in the vasculature of the heart and tail. As another example of nonretinal melanopsin expression we report here the previously unrecognized localization of this photopigment in nerve fibers within the cornea. Surprisingly, we were unable to detect light responses in the melanopsin-expressing corneal fibers in spite of our histological evidence based on genetically driven markers and antibody staining. We tested further for melanopsin localization in cell bodies of the trigeminal ganglia (TG), the principal nuclei of the peripheral nervous system that project sensory fibers to the cornea, and found expression of melanopsin mRNA in a subset of TG neurons. However, neither electrophysiological recordings nor calcium imaging revealed any light responsiveness in the melanopsin positive TG neurons. Given that we found no light-evoked activation of melanopsin-expressing fibers in cornea or in cell bodies in the TG, we propose that melanopsin protein might serve other sensory functions in the cornea. One justification for this idea is that melanopsin expressed in Drosophila photoreceptors can serve as a temperature sensor.
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http://dx.doi.org/10.1017/S0952523817000359DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6203320PMC
January 2018

Functional and Morphologic Alterations in Mechanical, Polymodal, and Cold Sensory Nerve Fibers of the Cornea Following Photorefractive Keratectomy.

Invest Ophthalmol Vis Sci 2018 05;59(6):2281-2292

Instituto Universitario Fernández-Vega, Universidad de Oviedo & Fundación de Investigación Oftalmológica, Oviedo, Spain.

Purpose: To define the characteristics and time course of the morphologic and functional changes experienced by corneal sensory nerves after photorefractive keratectomy (PRK).

Methods: Unilateral corneal excimer laser photoablation was performed in 54 anesthetized 3- to 6-month-old mice; 11 naïve animals served as control. Mice were killed 0, 3, 7, 15, and 30 days after PRK. Excised eyes were placed in a recording chamber superfused at 34°C. Electrical nerve impulse activity of single sensory terminals was recorded with a micropipette applied onto the corneal surface. Spontaneous and stimulus-evoked (cold, heat, mechanical, and chemical stimuli) nerve terminal impulse (NTI) activity was analyzed. Corneas were fixed and stained with anti-β-Tubulin III antibody to measure nerve density and number of epithelial nerve penetration points of regenerating subbasal leashes.

Results: Nerve fibers and NTI activity were absent in the injured area between 0 and 7 days after PRK, when sparse regenerating nerve sprouts appear. On day 15, subbasal nerve density reached half the control value and abnormally responding cold-sensitive terminals were recorded inside the lesion. Thirty days after PRK, nerve density was almost restored, active cold thermoreceptors were abundant, and polymodal nociceptor activity first reappeared.

Conclusions: Morphologic regeneration of subbasal corneal nerves started shortly after PRK ablation and was substantially completed 30 days later. Functional recovery appears faster in cold terminals than polymodal terminals, possibly reflecting an incomplete damage of the more extensively branched cold-sensitive axon terminals. Evolution of postsurgical discomfort sensations quality may be associated with the variable regeneration pattern of each fiber type.
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http://dx.doi.org/10.1167/iovs.18-24007DOI Listing
May 2018

Morphological and functional changes in TRPM8-expressing corneal cold thermoreceptor neurons during aging and their impact on tearing in mice.

J Comp Neurol 2018 08 2;526(11):1859-1874. Epub 2018 May 2.

Instituto Universitario Fernández-Vega, Universidad de Oviedo & Fundación de Investigación Oftalmológica, Oviedo, Spain.

Morphological and functional alterations of peripheral somatosensory neurons during the aging process lead to a decline of somatosensory perception. Here, we analyze the changes occurring with aging in trigeminal ganglion (TG), TRPM8-expressing cold thermoreceptor neurons innervating the mouse cornea, which participate in the regulation of basal tearing and blinking and have been implicated in the pathogenesis of dry eye disease (DED). TG cell bodies and axonal branches were examined in a mouse line (TRPM8 -EYFP) expressing a fluorescent reporter. In 3 months old animals, about 50% of TG cold thermoreceptor neurons were intensely fluorescent, likely providing strongly fluorescent axons and complex corneal nerve terminals with ongoing activity at 34°C and low-threshold, robust responses to cooling. The remaining TRPM8 corneal axons were weakly fluorescent with nonbeaded axons, sparsely ramified nerve terminals, and exhibited a low-firing rate at 34°C, responding moderately to cooling pulses as do weakly fluorescent TG neurons. In aged (24 months) mice, the number of weakly fluorescent TG neurons was strikingly high while the morphology of TRPM8 corneal axons changed drastically; 89% were weakly fluorescent, unbranched, and often ending in the basal epithelium. Functionally, 72.5% of aged cold terminals responded as those of young animals, but 27.5% exhibited very low-background activity and abnormal responsiveness to cooling pulses. These morpho-functional changes develop in parallel with an enhancement of tear's basal flow and osmolarity, suggesting that the aberrant sensory inflow to the brain from impaired peripheral cold thermoreceptors contributes to age-induced abnormal tearing and to the high incidence of DED in elderly people.
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http://dx.doi.org/10.1002/cne.24454DOI Listing
August 2018

Inhibitory Effect of Amitriptyline on the Impulse Activity of Cold Thermoreceptor Terminals of Intact and Tear-Deficient Guinea Pig Corneas.

J Ocul Pharmacol Ther 2018 Jan/Feb;34(1-2):195-203. Epub 2017 Nov 29.

1 Instituto de Neurociencias, Universidad Miguel Hernandez-CSIC , Alicante, Spain .

Purpose: Chronic dryness of the ocular surface evokes sensitization of corneal cold-sensitive neurons through an increase of sodium currents and a decrease of potassium currents, leading to the unpleasant dryness and pain sensations typical of dry eye disease. Here, we explored the effects of amitriptyline, a voltage-gated Na channel blocker used for the treatment of depression and chronic pain, on nerve terminal impulse (NTI) activity of cold-sensitive nerve terminals recorded in intact and tear-deficient guinea pig corneas.

Methods: Main lachrymal gland was surgically removed in anesthetized guinea pigs to induce chronic tear deficiency. Four to 6 weeks afterward, animals were sacrificed and both corneas placed in a perfusion chamber superfused at 34°C. Thermal stimuli were induced by changing the solution temperature from 34°C to 20°C (cooling ramp) and from 34°C to 50°C (heating ramp). Spontaneous and stimulus-evoked NTIs of cold-sensitive nerve terminals were recorded before, during, and after perfusion with solutions containing amitriptyline at different concentrations (3-30 μM).

Results: Perfusion with amitriptyline inhibited irreversibly and in a concentration-dependent manner the spontaneous NTI activity of cold thermoreceptors of intact corneas. This effect was less evident in tear-deficient corneas. In addition, amitriptyline (10 μM) attenuated the maximal response to cooling ramps without changing cold threshold in intact but not in tear-deficient corneas. Only cold thermoreceptors with low cooling threshold values were sensitive to amitriptyline.

Conclusion: Amitriptyline effectively reduces the activity of cold thermoreceptors, although its efficacy is different in intact and tear-deficient corneas, which might be due to the changes induced by ocular dryness in the expression of the various voltage-gated Na channels responsible of the action potential generation and propagation.
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http://dx.doi.org/10.1089/jop.2017.0066DOI Listing
October 2019

The Effect of Tear Supplementation with 0.15% Preservative-Free Zinc-Hyaluronate on Ocular Surface Sensations in Patients with Dry Eye.

J Ocul Pharmacol Ther 2017 Jul/Aug;33(6):487-492. Epub 2017 Apr 4.

1 Department of Ophthalmology, Semmelweis University , Budapest, Hungary .

Purpose: To evaluate the effect of tear supplementation with preservative free 0.15% zinc-hyaluronate on ocular surface sensations and corneal sensitivity in dry eye patients.

Methods: Ocular surface sensations were assessed using the ocular surface disease index (OSDI) questionnaire and by recording ocular sensations during forced blinking in parallel with noninvasive tear film breakup time measurement in 20 eyes of 20 dry eye patients. Corneal sensitivity thresholds to selective stimulation of corneal mechano-, thermal- and chemical receptors were measured using the Belmonte gas esthesiometer. All baseline measurements were repeated after 1 month of treatment with 0.15% zinc-hyaluronate.

Results: After 1 month, a significant decrease in mean OSDI score (from 35.66 ± 12.36 to 15.03 ± 11.22; P < 0.001) and a significant improvement in tear film breakup time (from 3.83 ± 0.80 to 8.67 ± 4.50 s; P < 0.001) was observed compared to baseline. Sensory responses during the interblink period also significantly decreased after 1 month (P < 0.004). Corneal sensitivity thresholds to mechanical stimulation (90.61 ± 20.35 vs. 103.92 ± 17.97 mL/min; P < 0.025) and chemical stimulation (33.21 ± 0.51 vs. 33.58% ± 0.44% CO; P < 0.025) significantly increased after 1 month, however sensitivity thresholds to thermal stimulation remained unchanged compared to baseline (P > 0.05).

Conclusion: Prolonged use of 0.15% zinc-hyaluronate results in an improvement of tear film stability and a decrease of dry eye complaints. The decrease in corneal mechano-and polymodal receptor excitability suggests that zinc-hyaluronate helps to recover normal corneal sensitivity, and thus might have a beneficial additional effect on reducing ocular surface complaints in dry eye patients.
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http://dx.doi.org/10.1089/jop.2016.0194DOI Listing
December 2017

Functional Properties of Sensory Nerve Terminals of the Mouse Cornea.

Invest Ophthalmol Vis Sci 2017 01;58(1):404-415

Instituto Universitario Fernández-Vega, Universidad de Oviedo & Fundación de Investigación Oftalmológica, Oviedo, Spain 2Instituto de Neurociencias, Universidad Miguel Hernández-Consejo Superior de Investigaciones Cientificas, San Juan de Alicante, Spain.

Purpose: To define the firing properties of sensory nerve terminals innervating the adult mouse cornea in response to external stimuli of differing modality.

Methods: Extracellular electrical activity of single corneal sensory nerve terminals was recorded in excised eyes of C57BL/6J mice. Eyes were placed in a recording chamber and were continuously superfused with warm saline solution. Nerve terminal impulse (NTI) activity was recorded by means of a glass pipette (tip ∼ 50 μm), applied on the corneal surface. Nerve terminal impulse discharges were stored in a computer for offline analysis.

Results: Three functionally distinct populations of nerve terminals were identified in the mouse cornea. Pure mechanonociceptor terminals (9.5%) responded phasically and only to mechanical stimuli. Polymodal nociceptor terminals (41.1%) were tonically activated by heat and hyperosmolal solutions (850 mOsm·kg-1), mechanical force, and/or TRPV1 and TRPA1 agonists (capsaicin and allyl isothiocyanate [AITC], respectively). Cold-sensitive terminals (49.4%) responded to cooling. Approximately two-thirds of them fired continuously at 34°C and responded vigorously to small temperature reductions, being classified as high-background activity, low-threshold (HB-LT) cold thermoreceptor terminals. The remaining one-third exhibited very low ongoing activity at 34°C and responded weakly to intense cooling, being named low-background activity, high-threshold (LB-HT) cold thermoreceptor terminals.

Conclusions: The mouse cornea is innervated by trigeminal ganglion (TG) neurons that respond to the same stimulus modalities as corneal receptors of other mammalian species. Mechano- and polymodal endings underlie detection of mechanical and chemical noxious stimuli while HB-LT and LB-HT cold thermoreceptors appear to be responsible for basal and irritation-evoked tearing and blinking, respectively.
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http://dx.doi.org/10.1167/iovs.16-20033DOI Listing
January 2017

Lacosamide diminishes dryness-induced hyperexcitability of corneal cold sensitive nerve terminals.

Eur J Pharmacol 2016 Sep 3;787:2-8. Epub 2016 Jun 3.

Instituto de Neurociencias, Universidad Miguel Hernández - CSIC, San Juan de Alicante, Spain. Electronic address:

Lacosamide is an anti-epileptic drug that is also used for the treatment of painful diabetic neuropathy acting through voltage-gated sodium channels. The aim of this work was to evaluate the effects of acute application of lacosamide on the electrical activity of corneal cold nerve terminals in lacrimo-deficient guinea pigs. Four weeks after unilateral surgical removal of the main lachrimal gland in guinea pigs, corneas were excised and superfused in vitro at 34°C for extracellular electrophysiological recording of nerve terminal impulse activity of cold thermosensitive nerve terminals. The characteristics of the spontaneous and the stimulus-evoked (cooling ramps from 34°C to 15°C) activity before and in presence of lacosamide 100µM and lidocaine 100µM were compared. Cold nerve terminals (n=34) recorded from dry eye corneas showed significantly enhanced spontaneous activity (8.0±1.1 vs. 5.2±0.7imp/s; P<0.05) and cold response (21.2±1.7 vs. 16.8±1.3imp/s; P<0.05) as well as reduced cold threshold (1.5±0.1 vs. 2.8±0.2 Δ°C; P<0.05) to cooling ramps compared to terminals (n=58) from control animals. Both lacosamide and lidocaine decreased spontaneous activity and peak response to cooling ramps significantly (P<0.05). Temperature threshold was increased by the addition of lidocaine (P<0.05) but not lacosamide (P>0.05) to the irrigation fluid. In summary, the application of lacosamide results in a significant decrease of the augmented spontaneous activity and responsiveness to cold of corneal sensory nerves from tear-deficient animals. Based on these promising results we speculate that lacosamide might be used to reduce the hyperexcitability of corneal cold receptors caused by prolonged ocular surface dryness due to hyposecretory or evaporative dry eye disease.
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http://dx.doi.org/10.1016/j.ejphar.2016.05.044DOI Listing
September 2016

Abnormal activity of corneal cold thermoreceptors underlies the unpleasant sensations in dry eye disease.

Pain 2016 Feb;157(2):399-417

Instituto de Neurociencias, Universidad Miguel Hernández-CSIC, San Juan de Alicante, Spain Department of Ophthalmology, Semmelweis University, Budapest, Hungary Laboratory of Neurophysiology, Department of Biomedicine, School of Medicine, University of Barcelona, Barcelona, Spain Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Barcelona, Spain Instituto Universitario Fernández-Vega, Universidad de Oviedo and Fundación de Investigación Oftalmológica, Oviedo, Spain Departamento de Fisiología, Genética y Microbiología, Universidad de Alicante, San Vicente del Raspeig, Spain.

Dry eye disease (DED) affects >10% of the population worldwide, and it provokes an unpleasant sensation of ocular dryness, whose underlying neural mechanisms remain unknown. Removal of the main lachrymal gland in guinea pigs caused long-term reduction of basal tearing accompanied by changes in the architecture and density of subbasal corneal nerves and epithelial terminals. After 4 weeks, ongoing impulse activity and responses to cooling of corneal cold thermoreceptor endings were enhanced. Menthol (200 μM) first excited and then inactivated this augmented spontaneous and cold-evoked activity. Comparatively, corneal polymodal nociceptors of tear-deficient eyes remained silent and exhibited only a mild sensitization to acidic stimulation, whereas mechanonociceptors were not affected. Dryness-induced changes in peripheral cold thermoreceptor responsiveness developed in parallel with a progressive excitability enhancement of corneal cold trigeminal ganglion neurons, primarily due to an increase of sodium currents and a decrease of potassium currents. In corneal polymodal nociceptor neurons, sodium currents were enhanced whereas potassium currents remain unaltered. In healthy humans, exposure of the eye surface to menthol vapors or to cold air currents evoked unpleasant sensations accompanied by increased blinking frequency that we attributed to cold thermoreceptor stimulation. Notably, stimulation with menthol reduced the ongoing background discomfort of patients with DED, conceivably due to use-dependent inactivation of cold thermoreceptors. Together, these data indicate that cold thermoreceptors contribute importantly to the detection and signaling of ocular surface wetness, and develop under chronic eye dryness conditions an injury-evoked neuropathic firing that seems to underlie the unpleasant sensations experienced by patients with DED.
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http://dx.doi.org/10.1097/j.pain.0000000000000455DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4733818PMC
February 2016

Corneal Sensitivity and Dry Eye Symptoms in Patients with Keratoconus.

PLoS One 2015 23;10(10):e0141621. Epub 2015 Oct 23.

Semmelweis University, Department of Ophthalmology, Budapest, Hungary.

Purpose: To investigate corneal sensitivity to selective mechanical, chemical, and thermal stimulation and to evaluate their relation to dry eye symptoms in patients with keratoconus.

Methods: Corneal sensitivity to mechanical, chemical, and thermal thresholds were determined using a gas esthesiometer in 19 patients with keratoconus (KC group) and in 20 age-matched healthy subjects (control group). Tear film dynamics was assessed by Schirmer I test and by the non-invasive tear film breakup time (NI-BUT). All eyes were examined with a rotating Scheimpflug camera to assess keratoconus severity.

Results: KC patients had significatly decreased tear secretion and significantly higher ocular surface disease index (OSDI) scores compared to controls (5.3±2.2 vs. 13.2±2.0 mm and 26.8±15.8 vs. 8.1±2.3; p<0.001). There was no significant difference in NI-BUT between the two groups (KC: 9.8±4.8 vs. control: 10.7±3.8; p>0.05). The mean threshold for selective mechanical (KC: 139.2±25.8 vs. control: 109.1±24.0 ml/min), chemical (KC: 39.4±3.9 vs. control: 35.2±1.9%CO2), heat (KC: 0.91±0.32 vs. control: 0.54±0.26 Δ°C) and cold (KC: 1.28±0.27 vs. control: 0.98±0.25 Δ°C) stimulation in the KC patients were significantly higher than in the control subjects (p<0.001, for all parameters). No correlation was found between age and mechanical, chemical, heat or cold thresholds in the patients with KC (p>0.05), whereas in the control subjects both mechanical (r = 0.52, p = 0.02), chemical (r = 0.47, p = 0.04), heat (r = 0.26, p = 0.04) and cold threshold (r = 0.40, p = 0.03) increased with age. In the KC group, neither corneal thickness nor tear flow, NI-BUT or OSDI correlated significantly with mechanical, chemical, heat or cold thresholds (p>0.05 for all variables).

Conclusions: Corneal sensitivity to different types of stimuli is decreased in patients with keratoconus independently of age and disease severity. The reduction of the sensory input from corneal nerves may contribute to the onset of unpleasant sensations in these patients and might lead to the impaired tear film dynamics.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0141621PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4619831PMC
June 2016

The Effect of Tear Supplementation on Ocular Surface Sensations during the Interblink Interval in Patients with Dry Eye.

PLoS One 2015 24;10(8):e0135629. Epub 2015 Aug 24.

Department of Ophthalmology, Semmelweis University, Budapest, Hungary.

Purpose: To investigate the characteristics of ocular surface sensations and corneal sensitivity during the interblink interval before and after tear supplementation in dry eye patients.

Methods: Twenty subjects (41.88±14.37 years) with dry eye symptoms were included in the dry eye group. Fourteen subjects (39.13±11.27 years) without any clinical signs and/or symptoms of dry eye were included in the control group. Tear film dynamics was assessed by non-invasive tear film breakup time (NI-BUT) in parallel with continuous recordings of ocular sensations during forced blinking. Corneal sensitivity to selective stimulation of corneal mechano-, cold and chemical receptors was assessed using a gas esthesiometer. All the measurements were made before and 5 min after saline and hydroxypropyl-guar (HP-guar) drops.

Results: In dry eye patients the intensity of irritation increased rapidly after the last blink during forced blinking, while in controls there was no alteration in the intensity during the first 10 sec followed by an exponential increase. Irritation scores were significantly higher in dry eye patients throughout the entire interblink interval compared to controls (p<0.004). NI-BUT significantly increased after HP-guar (p = 0.003) but not after saline drops (p = 0.14). In both groups, either after saline or HP-guar the shape of symptom intensity curves remained the same with significantly lower irritation scores (p<0.004), however after HP-guar the decrease was significantly more pronounced (p<0.004). Corneal sensitivity to selective mechanical, cold and chemical stimulation decreased significantly in both groups after HP-guar (p<0.05), but not after saline drops (p>0.05).

Conclusion: Ocular surface irritation responses due to tear film drying are considerably increased in dry eye patients compared to normal subjects. Although tear supplementation improves the protective tear film layer, and thus reduce unpleasant sensory responses, the rapid rise in discomfort is still maintained and might be responsible for the remaining complaints of dry eye patients despite the treatment.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0135629PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4547761PMC
May 2016

What Causes Eye Pain?

Curr Ophthalmol Rep 2015;3(2):111-121

Instituto de Neurociencias, Universidad Miguel Hernández-CSIC San Juan de Alicante, Avenida de la Universidad, s/n, 03202 Alicante, Spain.

Eye pain is an unpleasant sensory and emotional experience including sensory-discriminative, emotional, cognitive, and behavioral components and supported by distinct, interconnected peripheral and central nervous system elements. Normal or physiological pain results of the stimulation by noxious stimuli of sensory axons of trigeminal ganglion (TG) neurons innervating the eye. These are functionally heterogeneous. Mechano-nociceptors are only excited by noxious mechanical forces. Polymodal nociceptors also respond to heat, exogenous irritants, and endogenous inflammatory mediators, whereas cold thermoreceptors detect moderate temperature changes. Their distinct sensitivity to stimulating forces is determined by the expression of specific classes of ion channels: Piezo2 for mechanical forces, TRPV1 and TRPA1 for heat and chemical agents, and TRPM8 for cold. Pricking pain is evoked by mechano-nociceptors, while polymodal nociceptors are responsible of burning and stinging eye pain; sensations of dryness appear to be mainly evoked by cold thermoreceptors. Mediators released by local inflammation, increase the excitability of eye polymodal nociceptors causing their sensitization and the augmented pain sensations. During chronic inflammation, additional, long-lasting changes in the expression and function of stimulus-transducing and voltage-sensitive ion channels develop, thereby altering polymodal terminal's excitability and evoking chronic inflammatory pain. When trauma, infections, or metabolic processes directly damage eye nerve terminals, these display aberrant impulse firing due to an abnormal expression of transducing and excitability-modulating ion channels. This malfunction evokes 'neuropathic pain' which may also result from abnormal function of higher brain structures where ocular TG neurons project. Eye diseases or ocular surface surgery cause different levels of inflammation and/or nerve injury, which in turn activate sensory fibers of the eye in a variable degree. When inflammation dominates (allergic or actinic kerato-conjunctivitis), polymodal nociceptors are primarily stimulated and sensitized, causing pain. In uncomplicated photorefractive surgery and moderate dry eye, cold thermoreceptors appear to be mainly affected, evoking predominant sensations of unpleasant dryness.
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http://dx.doi.org/10.1007/s40135-015-0073-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4432221PMC
January 2015

Acid-sensing ion channels detect moderate acidifications to induce ocular pain.

Pain 2015 Mar;156(3):483-495

Neurophysiology Laboratory, Department of Physiological Sciences I, Medical School, Universitat de Barcelona, Barcelona, Spain Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Barcelona, Spain Department of Ophthalmology, Hospital Vall d'Hebron, Barcelona, Spain Instituto de Neurociencias, Universidad Miguel Hernandez-CSIC, San Juan de Alicante, Spain.

Sensory nerve fibers innervating the ocular anterior surface detect external stimuli producing innocuous and painful sensations. Protons are among the first mediators released by damaged cells during inflammation, tissue injury, or other chronic ophthalmic conditions. We studied whether acid-sensing ion channels (ASICs) are expressed in corneal sensory neurons and their roles in the response to moderate acidifications of the ocular surface and in pathologies producing ocular surface inflammation. Moderate acidic pH (6.6) activated ASIC-like currents in corneal sensory neurons, which were blocked by ASIC1- or ASIC3-specific toxins. Acidic pH depolarizes corneal sensory neurons to fire action potentials, an effect blocked by the ASIC3 inhibitor APETx2. 2-Guanidino-4-methylquinazoline, an ASIC3 agonist, activated a population of corneal polymodal sensory nerve fibers and significantly increased the blinking and tearing rate. The nocifensive behaviors produced by application of either a moderate acidic stimulus or ophthalmic drugs formulated in acidic solution were abolished by ASIC blockers. In a model of allergic keratoconjunctivitis, nocifensive behavior was greatly reduced by ASIC3 blockade, presumably by reducing nociceptor sensitization during the inflammatory process. Our results show that, in addition to the established role of TRPV1, ASICs play a significant role in the detection of acidic insults at the ocular surface. The identification of ASICs in corneal neurons and their alterations during different diseases is critical for the understanding of sensory ocular pathophysiology. They are likely to mediate some of the discomfort sensations accompanying several ophthalmic formulations and may represent novel targets for the development of new therapeutics for ocular pathologies.
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http://dx.doi.org/10.1097/01.j.pain.0000460335.49525.17DOI Listing
March 2015

Preclinical pharmacology, ocular tolerability and ocular hypotensive efficacy of a novel non-peptide bradykinin mimetic small molecule.

Exp Eye Res 2014 Nov 11;128:170-80. Epub 2014 Oct 11.

Institute of Neuroscience, University Miguel Hernandez-CSIC, San Juan Campus, Alicante, Spain.

We sought to characterize the ocular pharmacology, tolerability and intraocular pressure (IOP)-lowering efficacy of FR-190997, a non-peptidic bradykinin (BK) B2-receptor agonist. FR-190997 possessed a relatively high receptor binding affinity (Ki = 27 nM) and a high in vitro potency (EC50 = 18.3 ± 4.4 nM) for inositol-1-phosphate generation via human cloned B2-receptors expressed in host cells with mimimal activity at B1-receptors. It also mobilized intracellular Ca2+ in isolated human trabecular meshwork (h-TM), ciliary muscle (h-CM), and in immortalized non-pigmented ciliary epithelial (h-iNPE) cells (EC50s = 167-384 nM; Emax = 32-86% of BK-induced response). HOE-140, a selective B2-receptor antagonist, potently blocked the latter effects of FR-190997 (e.g., IC50 = 7.3 ± 0.6 nM in h-CM cells). FR-190997 also stimulated the release of prostaglandins (PGs) from h-TM and h-CM cells (EC50s = 60-84 nM; Emax = 29-44% relative to max. BK-induced effects). FR-190997 (0.3-300 μg t.o.) did not activate cat corneal polymodal nociceptors and did not cause ocular discomfort in Dutch-Belted rabbits, but it was not well tolerated in New Zealand albino rabbits and Hartley guinea pigs. A single topical ocular (t.o.) dose of 1% FR-190997 in Dutch-Belted rabbits and mixed breed cats did not lower IOP. However, FR-190997 efficaciously lowered IOP of conscious ocular hypertensive cynomolgus monkey eyes (e.g., 34.5 ± 7.5% decrease; 6 h post-dose of 30 μg t.o.; n = 8). Thus, FR-190997 is an unexampled efficacious ocular hypotensive B2-receptor non-peptide BK agonist that activates multiple signaling pathways to cause IOP reduction.
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http://dx.doi.org/10.1016/j.exer.2014.10.008DOI Listing
November 2014

Corneal sensory nerve activity in an experimental model of UV keratitis.

Invest Ophthalmol Vis Sci 2014 May 1;55(6):3403-12. Epub 2014 May 1.

Instituto de Neurociencias, Universidad Miguel Hernández-CSIC, San Juan de Alicante, Spain.

Purpose: To produce in guinea pigs a UV-induced keratitis, to analyze the effects of this pathology on corneal nerve activity.

Methods: In anesthetized animals, one eye was exposed to 254 nm UV-C radiation (500-1000 mJ/cm(2)), excised 24 to 48 hours later and superfused in vitro. Nerve impulse activity was recorded in ciliary nerve filaments or in corneal sensory terminals of intact and UV-irradiated eyes. Impulse activity in response to mechanical (von Frey hairs), chemical (98.5% CO2 gas jets), and thermal stimulation (cooling from 34°C to 20°C; heating to 50°C) was analyzed. Duration of eyelid closure and blinking and tearing rates were evaluated in control and in UV-irradiated eyes, before and after application of TRPV1, TRPA1, and TRPM8 agonists (100 μM capsaicin; 10 mM AITC, and 200 μM menthol, respectively).

Results: After irradiation, mechanical threshold of mechano-nociceptor corneo-scleral fibers was reduced (0.59 ± 0.4 vs. 0.27 ± 0.07 mN; P < 0.05) while polymodal nociceptors increased their response to chemical stimulation (1.7 ± 0.2 vs. 3.4 ± 0.5 imps/s; P < 0.05). In contrast, cold thermoreceptors showed a significantly lower ongoing activity at 34°C (8.6 ± 0.5 vs. 6.1 ± 0.9 imp/s; P < 0.05) and a reduced responsiveness to cooling pulses (peak frequency = 29.8 ± 1.3 vs. 18.9 ± 1.8 imp/s; P < 0.001). Blinking but not tearing rate was significantly higher; behavioral responses to topical capsaicin and AITC, but not to menthol were enhanced in UV-irradiated animals.

Conclusions: Sensitization of nociceptor and depression of cold thermoreceptor activity following UV radiation appear to result from an action of inflammatory mediators on TRP channels selectively expressed by sensory nerve terminals. Changes in nerve activity possibly underlie discomfort sensations associated with corneo-conjunctival inflammation induced by UV exposure.
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http://dx.doi.org/10.1167/iovs.13-13774DOI Listing
May 2014

Tear fluid hyperosmolality increases nerve impulse activity of cold thermoreceptor endings of the cornea.

Pain 2014 Aug 28;155(8):1481-1491. Epub 2014 Apr 28.

Instituto de Neurociencias, Universidad Miguel Hernandez-CSIC, San Juan de Alicante, Spain Fundacion de Investigación Oftalmológica, Instituto de Oftalmología Fernández-Vega, Oviedo, Spain.

Dry eye disease (DED) is a multifactorial disorder affecting the composition and volume of tears. DED causes ocular surface dryness, cooling, and hyperosmolality, leading ultimately to corneal epithelium damage and reduced visual performance. Ocular discomfort is the main clinical symptom in DED. However, the peripheral neural source of such unpleasant sensations is still unclear. We analyzed in excised, superfused mouse eyes, the effect of NaCl-induced hyperosmolality (325-1005 mOsm·kg(-1)) on corneal cold thermoreceptor and polymodal nociceptor nerve terminal impulse (NTI) activity. Osmolality elevations at basal corneal temperature (33.6°C) linearly increased the ongoing NTI frequency of cold thermoreceptors, at a mean rate of 0.34 imp·s(-1)/10 mOsm. This frequency increase became significant with osmolality values greater than 340 mOsm. Comparison of cold thermoreceptor activity increase induced by a dynamic temperature reduction of 1.8°C under iso- and hyperosmolal (360-mOsm) conditions provided evidence that more than 50% of the increased firing response was attributable to hyperosmolality. Comparatively, activation of corneal polymodal nociceptor endings by hyperosmolal solutions started with values of 600 mOsm and greater. Sensitization of polymodal nociceptors by continuous perfusion with an "inflammatory soup" (bradykinin, histamine, prostaglandin E2 [PGE2], serotonin, and adenosine triphosphate [ATP]) did not enhance their activation by hyperosmolal solutions. High osmolality also altered the firing pattern and shape of cold and polymodal NTIs, possibly reflecting disturbances in local membrane currents. Results strongly suggest that tear osmolality elevations in the range observed in DED predominantly excite cold thermoreceptors, supporting the hypothesis that dryness sensations experienced by these patients are due, at least in part, to an augmented activity of corneal cold thermoreceptors.
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http://dx.doi.org/10.1016/j.pain.2014.04.025DOI Listing
August 2014

Expression of cholecystokinin, gastrin, and their receptors in the mouse cornea.

Invest Ophthalmol Vis Sci 2014 Mar 28;55(3):1965-75. Epub 2014 Mar 28.

Fundación de Investigación Oftalmológica, Instituto Fernandez-Vega, Oviedo, Spain.

Purpose: Cholecystokinin (CCK) is a neuropeptide that has been identified in trigeminal ganglion neurons. Gastrin (GAST) is a related peptide never explored in the cornea. The presence and role of both gastrointestinal peptides in the cornea and corneal sensory neurons remain to be established. We explored here in mice whether CCK, GAST, and their receptors CCK1R and CCK2R are expressed in the corneal epithelium and trigeminal ganglion neurons innervating the cornea.

Methods: We used RT-PCR analysis to detect mRNAs of CCK, GAST, CCK1R, and CCK2R in mouse cornea epithelium, trigeminal ganglia, and primary cultured corneal epithelial cells. Immunofluorescence microscopy was used to localize these peptides and their receptors in the cornea, cultured corneal epithelial cells, and corneal nerves, as well as in the cell bodies of corneal trigeminal ganglion neurons identified by retrograde labeling with Fast Blue.

Results: Mouse corneal epithelial cells in the cornea in situ and in cell cultures expressed CCK and GAST. Only the receptor CCK2R was found in the corneal epithelium. In addition, mouse corneal afferent sensory neurons expressed CCK and GAST, and the CCK1R receptors.

Conclusions: The presence of CCK, GAST, and their receptors in the mouse corneal epithelium, and in trigeminal ganglion neurons supplying sensory innervation to the cornea, opens the possibility that these neuropeptides are involved in corneal neurogenic inflammation and in the modulation of repairing/remodeling processes following corneal injury.
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http://dx.doi.org/10.1167/iovs.13-12068DOI Listing
March 2014

The TFOS International Workshop on Contact Lens Discomfort: report of the subcommittee on neurobiology.

Invest Ophthalmol Vis Sci 2013 Oct 18;54(11):TFOS71-97. Epub 2013 Oct 18.

School of Optometry and Vision Science, University of New South Wales, Sydney, Australia.

This report characterizes the neurobiology of the ocular surface and highlights relevant mechanisms that may underpin contact lens-related discomfort. While there is limited evidence for the mechanisms involved in contact lens-related discomfort, neurobiological mechanisms in dry eye disease, the inflammatory pathway, the effect of hyperosmolarity on ocular surface nociceptors, and subsequent sensory processing of ocular pain and discomfort have been at least partly elucidated and are presented herein to provide insight in this new arena. The stimulus to the ocular surface from a contact lens is likely to be complex and multifactorial, including components of osmolarity, solution effects, desiccation, thermal effects, inflammation, friction, and mechanical stimulation. Sensory input will arise from stimulation of the lid margin, palpebral and bulbar conjunctiva, and the cornea.
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http://dx.doi.org/10.1167/iovs.13-13226DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5963174PMC
October 2013

Changes in sensory activity of ocular surface sensory nerves during allergic keratoconjunctivitis.

Pain 2013 Nov 16;154(11):2353-2362. Epub 2013 Jul 16.

Instituto de Neurociencias, Universidad Miguel Hernández-CSIC, Avda. Santiago Ramón y Cajal 2, E-03550 San Juan de Alicante, Spain.

Peripheral neural mechanisms underlying the sensations of irritation, discomfort, and itch accompanying the eye allergic response have not been hitherto analyzed. We explored this question recording the changes in the electrical activity of corneoconjunctival sensory nerve fibers of the guinea pig after an ocular allergic challenge. Sensitization was produced by i.p. ovalbumin followed by repeated application in the eye of 10% ovalbumin on days 14 to 18. Blinking and tearing rate were measured. Spontaneous and stimulus-evoked (mechanical, thermal, chemical) impulse activity was recorded from mechanonociceptor, polymodal nociceptor and cold corneoscleral sensory afferent fibers. After a single (day 14) or repeated daily exposures to the allergen during the following 3 to 4days, tearing and blinking rate increased significantly. Also, sensitization was observed in mechanonociceptors (transient reduction of mechanical threshold only on day 14) and in polymodal nociceptors (sustained enhancement of the impulse response to acidic stimulation). In contrast, cold thermoreceptors showed a significant decrease in basal ongoing activity and in the response to cooling. Treatment with the TRPV1 and TRPA1 blockers capsazepine and HC-030031 reversed the augmented blinking. Only capsazepine attenuated tearing rate increase and sensitization of the polymodal nociceptors response to CO2. Capsazepine also prevented the decrease in cold thermoreceptor activity caused by the allergic challenge. We conclude that changes in nerve impulse activity accompanying the ocular allergic response, primarily mediated by activation of nociceptor's TRPV1 and to a lesser degree by activation of TRPA1 channels, explain the eye discomfort sensations accompanying allergic episodes.
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http://dx.doi.org/10.1016/j.pain.2013.07.012DOI Listing
November 2013

Regenerative approaches as alternatives to donor allografting for restoration of corneal function.

Ocul Surf 2012 Jul 9;10(3):170-83. Epub 2012 May 9.

Integrative Regenerative Medicine Centre, Department of Clinical and Experimental Medicine, Linköping University, Sweden.

A range of alternatives to human donor tissue for corneal transplantation are being developed to address the shortfall of good quality tissues as well as the clinical conditions for which allografting is contraindicated. Classical keratoprostheses, commonly referred to as artificial corneas, are being used clinically to replace minimal corneal function. However, they are used only as last resorts, as they are associated with significant complications, such as extrusion/rejection, glaucoma, and retinal detachment. The past few years have seen significant developments in technologies designed to replace part or the full thickness of damaged or diseased corneas with materials that encourage regeneration to different extents. This review describes selected examples of these corneal substitutes, which range from cell-based regenerative strategies to keratoprostheses with regenerative capabilities via tissue-engineered scaffolds pre-seeded with stem cells. It is unlikely that one corneal substitute will be best for all indications, but taken together, the various approaches may soon be able to supplement the supply of human donor corneas for transplantation or allow restoration of diseased or damaged corneas that cannot be treated by currently available techniques.
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http://dx.doi.org/10.1016/j.jtos.2012.04.004DOI Listing
July 2012

Cold thermoreceptors, unexpected players in tear production and ocular dryness sensations.

Invest Ophthalmol Vis Sci 2011 Jun 1;52(6):3888-92. Epub 2011 Jun 1.

Instituto de Neurociencias de Alicante, Universidad Miguel Hernández-Consejo Superior de Investigaciones Científicas, San Juan de Alicante, Spain.

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http://dx.doi.org/10.1167/iovs.09-5119DOI Listing
June 2011

Corneal sensitivity in diabetic patients subjected to retinal laser photocoagulation.

Invest Ophthalmol Vis Sci 2011 Jul 29;52(8):6043-9. Epub 2011 Jul 29.

Instituto de Neurociencias de Alicante, Universidad Miguel Hernández-CSIC, Sant Joan d'Alacant, Spain.

Purpose: To determine the changes in corneal sensitivity to different stimulus modalities in diabetes mellitus (DM)1 and DM2 patients with retinopathy, and to explore whether argon laser photocoagulation exacerbates sensitivity loss in diabetic patients.

Methods: Corneal sensitivity to different modalities of stimulus was determined in one randomized eye in 52 patients with DM1 (n = 35) or DM2 (n = 17), and in 27 healthy subjects. Medical history was obtained from all the patients, including age, sex, time from DM diagnosis, type of diabetes, time from onset of retinopathy, type of diabetic retinopathy, and type of argon laser treatment. Corneal sensitivity was determined using a gas esthesiometer. Mechanical, chemical, and thermal (heat and cold) stimuli were applied on the central cornea.

Results: Sensitivity thresholds to selective mechanical, chemical, and cold stimulation were significantly higher in DM patients compared to controls. Sensitivity threshold to mechanical and chemical stimuli was higher in DM2 than in DM1 patients. In DM1 patients, mechanical threshold increased with time after DM diagnosis. No correlation was found between sensitivity thresholds to chemical or thermal stimulation and the age of the patient, type of retinopathy, or time from its diagnosis. Laser treatment generated a further impairment of corneal sensitivity.

Conclusions: Corneal sensitivity to mechanical, chemical, and thermal stimulation is decreased in DM patients, suggesting that diabetes affects homogeneously the different types of sensory neurons innervating the cornea. Corneal sensitivity appears to be more disturbed in DM2 than in DM1. Laser treatment of DM patients generates a further impairment in corneal sensitivity, probably as the result of physical damage to ciliary nerves.
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http://dx.doi.org/10.1167/iovs.10-7054DOI Listing
July 2011
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