Begoña Fonseca

Nucleotides in the Eye: Focus on Functional Aspects and Therapeutic Perspectives

The presence and activity of nucleotides and dinucleotides in the physiology of most, if not all, organisms, from bacteria to humans, have been recognized by the scientific community, and the eye is no exception. Nucleotides in the dynamic fluids interact with many ocular structures, such as the tears and aqueous humor. Moreover, high concentrations of nucleotides in these secretions may reflect disease states such as dry eye and glaucoma. Apart from the nucleotide concentration in these fluids, P2 purinergic receptors have been described on the ocular surface (cornea and conjunctiva), anterior pole (ciliary body, trabecular meshwork), and posterior pole (retina). P2X and P2Y purinergic receptors are essential in maintaining the homeostasis of ocular processes, such as tear secretion, aqueous humor production, or retinal modulation. When they are functioning properly, they allow the eye to do its job (to see), but in some cases, a lack or an excess of nucleotides or a malfunction in the corresponding purinergic receptors leads to disease. This Perspective is focused on the nucleotides and dinucleotides and the P2 purinergic receptors in the eye and how they contribute to normal and disease states. We also emphasize the action of nucleotides and their receptors and antagonists as potential therapeutic agents.

Melatonin analogue agomelatine reduces rabbit's intraocular pressure in normotensive and hypertensive conditions.

In the search for new compounds to reduce intraocular pressure (IOP), with fewer side effects, we have found that agomelatine, a melatonin analogue, can reduce IOP being, therefore, interesting for the treatment of ocular hypertension and glaucoma. In normotensive conditions, agomelatine (10μl 100μM) reduced IOP by 20.8±1.4% (n=18) with a maximal effect 180min after the compound application and 68.8±5.7% (n=8) in a hypertensive condition. Concentration-response curve depicted a sigmoid behaviour presenting a pD2 value of 9.7±0.3 which was equivalent to an EC50 of 0.19nM. The effect of agomelatine was partially antagonized by 4PPDOT (MT2 antagonist receptor. 10μl 100μM) and prazosin (MT3 antagonist receptor. 10μl 100μM) (85.6±1.6% and 87.2±1.9%, N=18 respectively.) Agomelatine hypotensive effect in normotensive condition was comparable to latanoprost (40μl) and brimonidine (40μl) and it was no so effective as dorzolamide (40μl) or timolol (40μl). These results may suggest the use of this melatonin analogue for the treatment of those ocular conditions, which involve an abnormal raise of intraocular pressure.

Effect of Melatonin and 5-Methoxycarbonylamino-N-Acetyltryptamine on the Intraocular Pressure of Normal and Glaucomatous Mice

Melatonin is a neurohormone that is produced not only by the pineal gland but also by several ocular structures. One of the main physiologic roles of melatonin is the reduction of intraocular pressure (IOP). Using both control C57BL/6J and glaucomatous DBA/2J mice as well as TonoLab tonometry, this study evaluated the effect of melatonin and 5-methoxycarbonylamino-N-acetyltryptamine (5-MCA-NAT) when glaucomatous pathology was fully established and compared pharmacological behavior in treated mice versus control mice. In addition, 5-MCA-NAT was tested to determine its effects on ameliorating increased IOP in a glaucoma model. The results demonstrate that melatonin and 5-MCA-NAT can reduce IOP in a concentration-dependent manner. The EC50 values for melatonin in control and glaucomatous animals were 34 µM and 50 µM, respectively. Interestingly, melatonin decreased IOP in 19.4% ± 3.7% and 32.6% ± 6.0% of control and glaucomatous mice, respectively, when the animals were studied at age 12 months. 5-MCA-NAT reduced IOP in the same manner and was able to stop IOP progression in glaucomatous mice. Use of melatonin receptor antagonists showed that hypotensive effects were blocked by the MT2 receptor antagonists luzindole and 4-phenyl-2-propionamidotetralin in the case of melatonin and by only 4-phenyl-2-propionamidotetralin in the case of 5-MCA-NAT. In conclusion, melatonin and 5-MCA-NAT can effectively reduce IOP in a glaucoma model, and their hypotensive effects are more profound in the glaucoma model than in control animals.

Dry Eye Treatment Based on Contact Lens Drug Delivery: A Review.

Abstract: Dry eye disease affects a substantial segment of the word population with increasing frequency. It is a multifactorial disease of the ocular surface and tear film, which causes ocular discomfort, visual disturbances, and tear instability with potential damage to the cornea and conjunctiva. Because of its multifactorial etiology, the use of different pharmacological treatment for dry eye treatment has been proposed, which include anti-inflammatory molecules, lubricants or comfort agents, and secretagogues. However, in some cases these pharmacological approaches only relieve symptoms temporarily, and consequently, eye care professionals continue to have difficulties managing dry eye. To improve pharmacological therapy that allows a more efficient and long-term action, effective ocular drug delivery of the currently available drugs for dry eye treatment is required. Contact lenses are emerging as alternative ophthalmic drugs delivery systems that provide an increased residence time of the drug at the eye, thus leading to enhanced bioavailability and more convenient and efficacious therapy. In this article, we reviewed the different techniques used to prepare contact lens-based drug delivery systems and focused on articles that describe the delivery of compounds for dry eye treatment through contact lenses.

Effect of overnight orthokeratology on conjunctival goblet cells

OBJECTIVE To evaluate the differences between goblet cell density (GCD) and symptomatology after one month of orthokeratology lens wear. METHODS A pilot, short-term study was conducted. Twenty-two subjects (29.7±7.0 years old) participated voluntarily in the study. Subjects were divided into two groups: habitual silicone hydrogel contact lens wearers (SiHCLW) and new contact lens wearers (NCLW). Schirmer test, tear break up time (TBUT), Ocular Surface Disease Index (OSDI) questionnaire and conjunctival impression cytology. GCD, mucin cloud height (MCH) and cell layer thickness (CLT) were measured. All measurements were performed before orthokeratology fitting and one month after fitting to assess the evolution of the changes throughout this time. RESULTS No differences in tear volume and TBUT between groups were found (p>0.05). However, the OSDI score was statistically better after one month of orthokeratology lens wear than the baseline for the SiHCLW group (p=0.03). Regarding the goblet cell analysis, no differences were found in CLT and MCH from the baseline visit to the one month visit for the SiHCLW compared with NCLW groups (p>0.05). At baseline, the GCD in the SiHCLW group were statistically lower than NCLW group (p<0.001). There was a significant increase in GCD after orthokeratology fitting from 121±140cell/mm(2) to 254±130cell/mm(2) (p<0.001) in the SiHCLW group. CONCLUSION Orthokeratology improves the dry eye subject symptoms and GCD after one month of wearing in SiHCLW. These results suggest that orthokeratology could be considered a good alternative for silicone hydrogel contact lens discomfort and dryness.

Changes in P2Y Purinergic Receptor Expression in the Ciliary Body in a Murine Model of Glaucoma

Glaucoma is a neuropathology, often accompanied by an elevated intraocular pressure (IOP), which can lead to blindness. Since DBA/2J mice develop glaucoma, several studies of the physiopathology of glaucoma have been reported in this animal model. It is also known that purinergic receptors are involved in the pathology of glaucoma by controlling aqueous humor production and drainage and therefore controlling IOP. There are no studies on purinergic receptors in the DBA/2J model of glaucoma and their relation to the development of the pathology, so the aim of this study was to make an approach to the purinergic mechanisms involved in glaucoma. All the experiments were performed using DBA/2J and C57BL/6J mice and investigating P2Y1, P2Y2, and P2Y6 receptors. IOP measurements were made with a non-invasive rebound tonometer, and animals were instilled with diadenosine tetraphosphate (Ap4A) and the corresponding purinergic antagonists in order to see their effects on IOP. The expression of mRNA for P2Y1, P2Y2, and P2Y6 purinergic receptors was carried out by quantitative real-time PCR. Additionally, P2Y-receptor expression was performed by immunohistochemical techniques carried out on the ciliary processes. The results showed that IOP decreases when Ap4A was instilled and that the expressions of the analyzed purinergic receptors were stable throughout all the ages under study in the C57BL/6J mice (control mice). On the other hand, there were significant changes in the purinergic receptor expression in DBA/2J suggesting that elevated IOP in these animals could be related to an increase of P2Y2 expression and a decrease in P2Y1 receptors.

Tau Triggers Tear Secretion by Interacting with Muscarinic Acetylcholine Receptors in New Zealand White Rabbits

In recent years, in vitro experiments have shown that the spread of Alzheimers disease is caused by a non-conventional activation of muscarinic receptors by dephosphorylated extracellular tau protein. However, so far, in vivo data to support this hypothesis has not been obtained. The eye provides a good model where cholinergic (muscarinic) transmission can be analyzed. The role of muscarinic receptors in the stimulation of lacrimal gland secretion has already been described, and it has been suggested that acetylcholine is the main transmitter controlling tear secretion. In this project, we have studied the interaction between tau and muscarinic receptors by analyzing tear secretion in the eyes of white rabbits. Our results show that tau protein increases tear secretion by 47.2% in a similar way to a muscarinic receptor agonist carbachol (84.3%). The use of muscarinic antagonists indicated that tau interacts with M1 and mainly M3 muscarinic receptors. In summary, tau can bind muscarinic receptors in vivo and this may explain the spread of the pathology.