Dry-responsive corneal afferents, TRPM8, and regulation of tears

描述(申请人提供)：干眼症(DES)是由于角膜表面泪膜不充分引起的。因此，DES患者遭受眼睛疼痛，在某些情况下还会出现严重的视力问题。DES影响多达20%的人口，治疗往往仍然不足。干眼的原因之一可能是感觉神经元无法通过对角膜干燥的反应来正确评估眼表液体状态。维持正常泪膜的神经调节机制，通过基础泪液的产生，目前尚不清楚。我们最近发现，一类角膜初级传入神经元，即冷细胞，已知对非伤害性降温有反应，也可以通过干燥眼表、高渗溶液和薄荷醇来激活。由于角膜表面的泪液蒸发会导致泪膜降温和渗透压增加，参与基底部撕裂调节的神经元应该具有这些特性。据推测，这些对干燥有反应的冷细胞代表反射弧的传入分支，它驱动非伤害性的基底部撕裂，而不会产生眼痛。这一假设将从四个具体目标进行检验。第一个目标将确定由眼表面干燥所激活的角膜初级传入神经元的编码特性。特别关注的是高渗和TRPM8依赖的诱发反应；TRPM8通道可以被薄荷醇和无害的降温激活。角膜初级传入将在大鼠三叉神经节中用在体单单位电生理学进行表征。本课程将检查对眼表面干燥的反应，以及由热、化学和渗透刺激引起的反应。目的2将确定TRPM8激动剂改变角膜初级传入神经元冷和干诱发反应的能力。目的3将确定干燥眼表面激活的三叉神经脊束核(VSP)神经元的编码特性和投射靶点。VSP中的单个单位记录将用于评估从角膜初级传入直接输入的神经元的特性，并将使用电刺激来测试其向参与调节撕裂的区域的投射状态。最后，目标4将确定三叉神经脊束核内的回路，参与大鼠眼表应用TRPM8和TRPA1激动剂引起的撕裂和伤害性行为。将对接受角膜输入的VSP的两个不同区域的病变进行检查，以确定它们对眼刺激的撕裂和无反应的相对贡献。增加对眼表面干燥反应的神经元的敏感性和活性的能力代表了潜在的DES治疗的新策略。

DESCRIPTION (provided by applicant): Dry eye syndrome (DES) results from an inadequate tear film on the corneal surface. As a result, patients with DES suffer from ocular pain and in some cases serious vision problems. DES affects as much as 20% of the population and treatment often remains inadequate. One cause of dry eye may be an inability of sensory neurons to properly assess the ocular surface fluid status by responding to drying of the cornea. The neural regulatory mechanisms involved in maintaining a normal tear-film, via basal tear production, are still unknown. We have recently discovered that a class of corneal primary afferent neurons, cold cells, which are known to respond to non-noxious cooling, are also activated by drying of the ocular surface, hyperosmotic solutions, and menthol. These properties would be expected of neurons that participate in the regulation of basal tearing, as evaporation of tears from the corneal surface causes both cooling and increased osmolarity of the tear film. It is hypothesized that these dry-responsive cold cells represent the afferent limb of the reflex arc that drives non- noxious basal tearing without producing ocular pain. This hypothesis will be examined in four specific aims. The first aim will determine the encoding properties of corneal primary afferent neurons activated by drying of the ocular surface. A particular focus will be on hyperosmotic and TRPM8 dependent evoked responses; TRPM8 channels are activated by both menthol and innocuous cooling. Corneal primary afferents will be characterized using in vivo single-unit electrophysiology in the rat trigeminal ganglion. Responses to drying of the ocular surface will be examined, as well as responses evoked by thermal, chemical, and osmotic stimuli. Aim 2 will determine the ability of TRPM8 agonists to alter cold- and dry-evoked responses in corneal primary afferent neurons. Aim 3 will determine the encoding properties and projection targets of spinal trigeminal nucleus (Vsp) neurons activated by drying of the ocular surface. Single unit recordings in Vsp will be used to assess properties of neurons that receive direct input from corneal primary afferents, and electrical stimulation will be employed to test for their projection status to regions involved in the regulation of tearing. Finally, Aim 4 will determine the circuitry within the spinal trigeminal nucleus involved in lacrimation and nocifensive behaviors evoked by TRPM8 and TRPA1 agonists applied to the ocular surface in rats. Lesions in two distinct regions of Vsp that receive corneal inputs will be performed to determine their relative contribution to tearing and nocifensive responses to ocular stimulation. The ability to increase the sensitivity and activity of neurons that respond to drying of the ocular surface represents a novel strategy for the potential treatment of DES.