Anatomic Variations of the Lateral Branch of the Supraorbital Nerve Observed in Endoscopic Forehead Surgery.

Surgeons dissect carefully in the medial third of the supraorbital rim to preserve the supraorbital nerve (SON) during surgical forehead rejuvenation. However, the anatomic variations of SON exit from the frontal bone have been researched in cadaver or imaging studies. In this study, we report a variation in the lateral branch of SON observed in an endoscopic view during forehead lifts. A retrospective review of 462 patients who underwent endoscopy-assisted forehead lifts between January 2013 and April 2020 was performed. Data, including the location, number, and form of the exit point and thickness of SON and its lateral branch variant, were recorded and reviewed intraoperatively, utilizing high-definition endoscopic assistance. Thirty-nine patients and 51 sides were included, and all patients were female, with a mean age of 44.53 (18-75) years. This nerve exited a foramen in the frontal bone ~8.82 ± 2.79 mm lateral to SON and ~1.89 ± 1.34 mm from the supraorbital margin vertically. Observed thickness variations of the lateral branch of SON included 20 small, 25 medium, and 6 large nerves. This study revealed various positional and morphologic variations of the lateral branch of SON in an endoscopic view. Thus, surgeons can be alerted of the anatomic variations of SON and establish careful dissection during procedures. In addition, the findings of this study will be useful in planning nerve blocks, filler injections, and migraine treatments in the supraorbital region.

Sensory Innervation of the Upper Eyelid and Its Implication for Upper Eyelid Surgery.

PURPOSE: Sensory alterations of the upper eyelid skin and eyelashes are frequently encountered after upper eyelid surgery. The objective of this study was to provide information on the exact course and distribution of sensory nerve fibers through the anatomic planes in the upper eyelid. METHODS: Ten formalin-fixed hemifaces were dissected. The nerve branches of the ophthalmic nerve in the upper eyelid were traced in an anterograde fashion. RESULTS: A total of 151 nerve fibers were recorded during dissection. The infratrochlear, supratrochlear, supraorbital, and lacrimal nerve contributed each to both the upper eyelid skin innervation and the upper eyelid rim plexus in different distribution patterns. The mean distance from the eyelid margin at which nerve fibers pierced from preseptal into the orbicularis muscle was 14 ± 1.1 mm for nerve fibers targeting the eyelid dermis and 3.7 ± 1.2 mm for nerve fibers targeting the eyelid rim plexus ( p < 0.001). The mean intraorbicular course of nerve fibers was 3 mm (0-17; standard deviation 4.1). The mean distance from the eyelid margin at which nerve fibers pierced from the orbicularis muscle into the preorbicular plane was 10 ± 1 mm for nerve fibers innervating the eyelid dermis and 1.3 ± 0.8 mm for nerve fibers innervating the eyelid rim plexus ( p < 0.001). The mean distance of the preorbicular course of nerve fibers was 2 mm (0-15; standard deviation 3.6). CONCLUSIONS: Based on the findings, a certain degree of postoperative eyelid skin numbness is inevitable while eyelash innervation may be spared in upper blepharoplasty.

How the Lifting Amount of Endoscopic Brow Lifts Is Influenced by Supraorbital Nerve Tension and Brow Gliding-Layer Mobility.

BACKGROUND: Sensory nerve tension and gliding-layer mobility in the brow may be significant factors affecting postoperative brow level in an endoscopic brow lift, yet they have rarely been studied. METHODS: To investigate the effects of sensory nerve tension and gliding-layer mobility, the following measurements were performed alongside the endoscopic brow lift in 50 fresh cadaveric hemifaces: amount of brow elevation, critical lifting amount (as sensory nerves became tense), laxity of sensory nerve courses, and mobility of brow-gliding layers. The sensory nerve situations in the subperiosteal and subgaleal dissections were also observed. RESULTS: Supraorbital nerve tension limited the cephalic advancement of the forehead flap. The mean elevation of the brow was 5.8 ± 1 mm (range, 3.5 to 8.6 mm). The mean critical lifting amount was 5.3 ± 1.1 mm (range, 4.0 to 7.3 mm). The mean amount of laxity in the supraorbital nerve (the permissible amount of lift) was 4.1 ± 0.9 mm (range, 2.5 to 5.5 mm). The galeal fat pad was responsible for 60% of brow mobility. The sensory nerve was more protected by a subgaleal dissection in the brow and inferior forehead and by a subperiosteal dissection in the middle and upper forehead. CONCLUSIONS: Cephalic movement of the forehead flap is limited by supraorbital nerve tension. The permitted lifting amount varies from 2.5 to 5.5 mm. Gliding-layer mobility in the brow offsets the postoperative amount of cephalic advancement of the forehead flap. Consideration of supraorbital nerve tension and gliding-layer mobility is recommended to obtain an optimal brow level in endoscopic brow lifts.

Central Corneal Subbasal Nerve Plexus Abnormalities in Sjögren Disease: A Pilot Study.

PURPOSE: Small-fiber neuropathy (SFN) is known to be associated with Sjögren disease (SjD), and in vivo corneal confocal microscopy can identify features compatible with SFN. Here, we performed a descriptive study to identify features of SFN of the corneal subbasal nerve plexus using in vivo confocal microscopy. METHODS: We recruited 10 participants from the Sjögren's International Collaborative Clinical Alliance (SICCA), 1 new participant (in an effort to expand the SICCA cohort), and 22 healthy controls. All participants underwent slit-lamp examination and in vivo confocal microscopy of the central corneal subbasal nerve plexus centered about the central whorl to create a 30-image montage. Each image was analyzed with automated software (ACCmetrics, Manchester, United Kingdom) to produce 7 nerve metrics. We performed t-tests and age-adjusted regressions to make comparisons of nerve metrics between participants with SjD and healthy controls. RESULTS: Most nerve metrics were significantly lower in participants with SjD compared with healthy controls. The mean corneal nerve fiber density was found to be 3.5 mm/mm 2 in participants with SjD compared with 10.6 mm/mm 2 in healthy controls (95% confidence interval, -8.4 to -0.93; P = 0.02). Within the 11 participants with SjD, 22 eyes were analyzed on confocal microscopy, and 16 of those eyes (from 9 individuals) did not have an identifiable central whorl. Within the 22 healthy controls, 22 eyes (right eye alone) were analyzed on confocal microscopy, and 21 of those eyes had an identifiable central whorl. CONCLUSIONS: SjD exhibits lower corneal nerve metrics compared with healthy controls. These findings suggest that features compatible with SFN can distinguish SjD from healthy controls and may serve as a potential novel biomarker in identifying SjD.

Investigation of anterior segment structures of the eye in Behçet's disease using in vivo confocal microscopy.

PURPOSE: We sought to investigate alterations in the corneal subbasal nerve plexus and endothelium in patients with Behçet's disease (BD). METHODS: This cross-sectional study included 64 patients with BD and 30 age- and gender-matched healthy control subjects. Those with BD were classified as having ocular or non-ocular disease. All subjects underwent a corneal endothelial and subbasal nerve density evaluation using in vivo confocal microscopy (IVCM). The differences among groups were analyzed using the Kruskal-Wallis test followed by Dunn's multiple comparison procedure. RESULTS: The mean age of study participants was 35.7 ± 10.2 years (16-58) in the ocular BD group, 39.6 ± 14.9 years (11-66) in the non-ocular BD group, and 34.1 ± 11.2 years (21-55) in the control group. No statistical significance was found in terms of age (p = 0.259) or sex (p = 0.560) between groups. The mean endothelial cell density determined with IVCM was 2124.9 [Formula: see text] 417.4 cells/mm2 (1811-3275) in the ocular group and 2546 [Formula: see text] 335 cells/mm2 (1798-3280) in the control group (p = 0.000). In the ocular group, the mean density of the subbasal nerve plexus was significantly lower (p = 0.004), and nerve tortuosity was significantly higher (p = 0.002). CONCLUSIONS: Ocular BD could be responsible for changes in the corneal layers, especially endothelial and corneal nerve structures. Nerve density and tortuosity differences could be inflammatory indicators for BD.

In vivo confocal microscopic study of corneal innervation in Sjögren's Syndrome with or without small fiber neuropathy.

PURPOSE: To study changes in the subbasal nerve plexus by In vivo confocal microscopy (IVCM) in Sjögren's Syndrome (SS) with or without associated Small Fiber Neuropathy (SFN), in order to prevent diagnostic delay. METHODS: Seventy-one patients with SS, including 19 with associated SFN, 20 healthy volunteers and 20 patients with Meibomian gland dysfunction (MGD) were included in this retrospective case-control study. IVCM was used to investigate subbasal nerve plexus density and morphology. RESULTS: Corneal sensitivity as evaluated with the Cochet-Bonnet aesthesiometer was significantly reduced in the SS group versus the control group (P = 0.026) and the MGD group (P = 0.037). The number of inflammatory cells was significantly increased in the SS group to 86.2 ± 82.1 cells/mm2 compared to the control group (P < 0.001). The density of the subbasal nerve plexus was significantly reduced to 16.7 ± 6.5 mm/mm2 in the SS group compared to the control group (P < 0.005) and the MGD group (P = 0.042). The tortuosity of the nerves in the SS group was significantly increased compared to the control group (P < 0.001) and the MGD group (P = 0.025). The average number of subbasal nerve plexus neuromas was significantly increased in the SS group compared to the control group (P = 0.001), with a significant increase in the average number of neuromas in SS patients with associated SFN compared to SS patients without SFN (P = 0.008). CONCLUSION: IVCM can be useful to detect corneal nerve changes in SS patients and may allow earlier diagnosis of the disease and to consider new therapeutic approaches.

Diurnal Variation of Corneal Dendritic Cell Density.

Purpose: To measure variation in corneal dendritic cell density, and percentage of mature to total dendritic cells, in healthy individuals during the sleep/wake cycle.Methods: Using in vivo confocal microscopy, images of the subbasal nerve plexus were captured from 19 healthy, noncontact lens wearing participants. The central cornea and inferior whorl were imaged three times (midday, before sleep, upon awakening). Dendritic cell counts from the images were categorized according to perceived maturity (immature vs mature). Dendritic cell density and percentage of mature to total cells were compared between time points.Result: The median and interquartile range (IQR) of total dendritic cell density in the central cornea was 32.0 (7.0-131.3) cells/mm2 at midday, 37.1 (8.2-103.9) cells/mm2 before sleep, and 19.5 (7.0-83.2) cells/mm2 on awakening. Corresponding values for immature cells were 28.1 (5.8-112.5) cells/mm2, 22.3 (7.4-84.0) cells/mm2 and 18.0 (2.9-64.8) cells/mm2, and for mature cells, 3.1 (0.0-6.6) cells/mm2, 2.0 (0.8-16.8) cells/mm2, and 1.6 (0.2-8.2) cells/mm2. At the inferior whorl, total dendritic cell density was 38.5 (18.4-84.5) cells/mm2, 34.4 (9.4-82.3) cell/mm2, and 32.3 (15.2-96.1) cells/mm2. Immature cell density was 32.8 (18.4-80.9) cells/mm2, 34.4 (8.6-81.0) cells/mm2, and 32.3 (12.6-78.5) cells/mm2. Mature cell density was 1.6 (0.0-6.3) cells/mm2, 1.6 (0.0-3.1) cells/mm2, and 1.8 (0.0-6.3) cells/mm2. There was no significant difference between time points for total cell density (p > 0.05), but the percentage of mature cells upon awakening was significantly greater, compared to midday, at the central cornea (p = 0.02).Conclusion: In healthy individuals, overall corneal dendritic cell density is reasonably constant during the sleep/wake cycle, but the relative number of mature cells tends to increase overnight.

Corneal Nerve Parameter Reference Values for Chinese Adults Assessed by Corneal Confocal Microscopy.

Background: Confocal corneal microscopy is an excellent new noninvasive tool for assessing diabetic peripheral neuropathy. We aimed to investigate the clinical variables associated with corneal nerve parameters and establish reference values for clinical use in healthy Chinese adults. Methods: The study enlisted 257 healthy volunteers (137 females and 120 males) from two clinical academic centers in China. Two experts captured and selected images of the central corneal subbasal nerve plexus at each center using the same corneal confocal microscopy instrument according to a commonly adopted protocol. Corneal nerve fiber density (CNFD), corneal nerve branch density (CNBD), and corneal nerve fiber length (CNFL) were measured using fully automated software (ACCMetrics). The correlation between clinical indicators and confocal corneal microscopy measures was determined using partial correlation. Quantile regression was used to calculate reference values and estimate the effects of clinical factors on the normative values of confocal corneal microscopy measures. Results: Females had significantly higher CNFD, CNBD, and CNFL than males. There was no correlation between age, glycated hemoglobin (HbA1c), height, weight, body mass index (BMI), and any corneal nerve fiber parameter in both sexes. In either sex, age, weight, height, BMI, and HbA1c did not affect the 0.05th quantile values of any corneal nerve parameter. Conclusions: This study establishes sex-adjusted reference values for corneal confocal microscopy measures in Chinese adults and provides a reference for clinical practice and research with this technique.

Corneal Sensation and Nerve Fiber Changes in Patients With Interstitial Cystitis.

PURPOSE: Central sensitization syndromes are associated with ocular surface discomfort and certain changes in corneal sensation. The aim of this study was to evaluate corneal changes in patients with interstitial cystitis (IC). METHODS: Thirty patients with IC and 32 healthy control subjects were included in this study. All patients had a detailed ophthalmological examination including measurement of corneal sensation with Cochet-Bonnet esthesiometer, tear breakup time, Schirmer I test, and Ocular Surface Disease Index questionnaire. After these examinations, corneal subbasal nerve plexus of the patients was evaluated with in vivo corneal confocal microscopy (IVCM) and the images were analyzed using fully automated software (ACC Metrics Corneal Nerve Fiber Analyzer V.2). RESULTS: There was no significant difference between the groups regarding age and gender distribution. Corneal sensation was significantly higher in patients with IC (P = 0.03), whereas tear breakup time, Schirmer I test, and Ocular Surface Disease Index scores were similar between the patients and controls. IVCM demonstrated nerve fiber loss in patients with IC. Corneal nerve fiber density, corneal nerve branch density, and corneal nerve fiber length were significantly reduced in patients with IC compared with healthy controls (P < 0.001, P = 0.04, and P < 0.001, respectively). CONCLUSIONS: Patients with IC had increased corneal sensation and decreased nerve fiber density in IVCM analysis. Corneal nerve fiber loss might decrease the induction threshold of the remaining fibers to induce peripheral sensitization, which may also trigger central sensitization in long term.

Structural and functional alterations in corneal nerves after single-step transPRK.

PURPOSE: To evaluate the relationship between structural and functional changes of corneal subbasal nerves after single-step transepithelial photorefractive keratectomy (TransPRK). SETTING: Tianjin Medical University Eye Hospital. DESIGN: Prospective observational study. METHODS: 51 eligible candidates who underwent TransPRK for moderate myopia were included. The regeneration of corneal subbasal nerves were evaluated by in vivo confocal microscopy (IVCM) prior to surgery and at 1 week and 1 month, 3 months, 6 months, and 12 months postoperatively. The corneal sensitivity was measured by Cochet-Bonnet esthesiometer. The number of complete and incomplete blinks were recorded by LipiView interferometer. A correlation analysis was performed between these variables. RESULTS: 51 eyes of 51 patients were enrolled in this study. IVCM revealed that the density and length of corneal nerve decreased immediately and did not return to preoperative levels by 12 months post-TransPRK ( P < .01). The mean corneal sensitivity was slightly lower compared with preoperative levels at 1 week postoperatively ( P = .001) and then increased to baseline preoperative level within 1 month postoperatively. Mean total blink frequency and the partial blink rates had no statistically significant change postoperatively ( P = .087). There was no direct correlation between postoperative recovery of corneal sensitivity, changes in blink frequency, and regeneration of subbasal corneal nerve. CONCLUSIONS: The significant and prolonged decrease in corneal subbasal nerve morphology was accompanied by only a slight and transient reduction in corneal sensitivity to mechanical stimuli within 1 month after TransPRK. The results demonstrated that the structural changes of subbasal nerves were not parallel to their functional changes after TransPRK.

Corneal Nerve Fiber and Sensitivity Loss After Repeated Intravitreal Anti-VEGF Injections: An In Vivo Confocal Microscopy Study.

PURPOSE: The purpose of this study was to investigate corneal sensation, subbasal nerve plexus (SBNP), and ocular surface symptoms in patients who underwent multiple intravitreal antivascular endothelial growth factor (anti-VEGF) injections for age-related macular degeneration (AMD) and diabetic macular edema (DME). METHODS: Forty patients with previous anti-VEGF intravitreal injections (20 AMD and 20 DME) and 30 healthy controls were included in this study. In vivo corneal confocal microscopy (IVCM) of the SBNP, corneal sensitivity measurement with a Cochet-Bonnet esthesiometer, noninvasive tear break-up times, and ocular surface disease score index (OSDI) calculation were performed for each participant. Corneal nerve fiber density, corneal nerve branch density, total length of all nerve fibers, corneal total branch density, corneal nerve fiber area, corneal nerve fiber width, and corneal nerve fiber fractal dimension parameters were obtained by automatic digital analysis. RESULTS: Corneal nerve fiber density, corneal nerve branch density, total length of all nerve fibers, and corneal nerve fiber fractal dimension in IVCM imaging and corneal sensitivity were significantly decreased in both AMD and DME groups compared with the control group. Corneal nerve fiber width and OSDI scores were significantly increased in AMD and DME groups compared with the control group. None of the IVCM parameters were significantly different between AMD and DME groups. Corneal sensitivity was decreased in patients with DME compared with patients with AMD. Tear break-up time was not different among the groups. CONCLUSIONS: Corneal SBNP parameters were affected, corneal sensitivity was decreased, and OSDI scores were increased in patients with multiple intravitreal anti-VEGF injections. IVCM parameters were not significantly different between AMD and DME groups.

Effect of herpes simplex keratitis scar location on bilateral corneal nerve alterations: an in vivo confocal microscopy study.

AIMS: To evaluate the impact of herpes simplex virus (HSV)-induced scar location on bilateral corneal nerve alterations using laser in vivo confocal microscopy (IVCM). METHODS: Central and peripheral corneal subbasal nerve density (CSND) were assessed bilaterally in 39 patients with unilateral HSV-induced corneal scars (21 central scars (CS), 18 peripheral scars (PS)) using IVCM. Results were compared between patients and 24 age-matched controls. CSND was correlated to corneal sensation for all locations. RESULTS: Overall patients revealed significant decrease of CSND in the central and peripheral cornea (9.13±0.98 and 6.26±0.53 mm/mm2, p<0.001), compared with controls (22.60±0.77 and 9.88±0.49 mm/mm2). CS group showed a decrease in central (8.09±1.30 mm/mm2) and total peripheral nerves (5.15±0.62 mm/mm2) of the affected eyes, whereas PS group demonstrated a decrease in central (10.34±1.48 mm/mm2) and localised peripheral nerves only in the scar area (4.22±0.77 mm/mm2) (all p<0.001). In contralateral eyes, CSND decreased in the central cornea of the CS group (16.88±1.27, p=0.004), and in the peripheral area, mirroring the scar area in the affected eyes of the PS group (7.20±0.87, p=0.032). Corneal sensation significantly decreased in the whole cornea of the affected, but not in contralateral eyes (p<0.001). A positive correlation between CSND and corneal sensation was found in all locations (p<0.001). CONCLUSIONS: Patients with HSV scar demonstrate bilateral CSND decrease as shown by IVCM. CSND and corneal sensation decrease in both central and peripheral cornea in affected eyes, although only in the scar area in PS group. Interestingly, diminishment of CSND was found locally in the contralateral eyes, corresponding and mirroring the scar location in the affected eyes.

Corneal sub-basal nerve plexus microneuromas in individuals with and without dry eye.

BACKGROUND/AIM: An objective marker is needed to detect when corneal nerve abnormalities underlie neuropathic corneal pain (NCP), as symptoms often overlap with those of dry eye (DE). This study evaluated microneuroma (MN) frequency in various populations and investigated relationships between MN presence and DE clinical features in individuals with DE symptoms but without a history of refractive surgery, in order to eliminate refractive surgery as a potential confounder of nerve abnormalities. METHODS: This was a retrospective study that included individuals with and without DE symptoms who underwent a clinical evaluation for DE (symptom surveys and ocular surface evaluation) and in vivo confocal microscopy imaging. DE clinical features (including those suggestive of neuropathic pain) were compared based on MN presence using t-tests, χ2 analyses and Pearson's correlation coefficients with 0.05 alpha level. RESULTS: MN frequencies did not significantly differ between individuals with DE symptoms (Dry Eye Questionnaire 5 score ≥6) and a history of refractive surgery (n=1/16, 6%), individuals with DE symptoms without a history of refractive surgery (n=26/119, 22%) and individuals without DE symptoms (n=2/18, 11%, p=0.22). Among individuals with DE symptoms without a history of refractive surgery, DE clinical features, including those indicative of NCP (burning sensation and sensitivity to light, wind and extreme temperatures), did not significantly differ based on MN presence (p>0.05). CONCLUSION: MN frequencies did not significantly differ between individuals with and without DE symptoms. Their presence alone could not distinguish between DE subtypes, including features of NCP in our study population.

Orbital neuromuscular choristoma of the ophthalmic nerve.

INTRODUCTION: Neuromuscular choristoma (NC) is a rare tumor composed of striated muscle fibers admixed with nerve fibers that often affects large peripheral nerve trunks in children. To the best of our knowledge, this is the first reported case in the literature of a NC of the ophthalmic nerve in an adult. CASE DESCRIPTION: A 27-year-old woman presented with a 20-day history of left periorbital headache radiating to the frontotemporal region, with associated tearing, nausea, and vomiting. The examination was normal, except for hypoesthesia in the distribution of the first trigeminal nerve branch (V1). Cranial computerized tomography showed a lesion in the left orbit and cavernous sinus with widening of the superior orbital fissure. On magnetic resonance imaging, the lesion was well-defined with fusiform morphology and showed a central cystic component and peripheral enhancement. An extradural approach was performed with subtotal tumor resection. Biopsy showed proliferation of striated muscle cells admixed with nerve fascicles and Schwann cells. A pathological diagnosis of NC of V1 was determined. Six months after surgery, left hypotropia with limitation of supraduction was observed, as well as hypoesthesia and paralysis of the left forehead with ipsilateral eyebrow ptosis. The rest of the examination remained within normal limits. CONCLUSION: NC should be considered in the differential diagnosis of a young patient with periorbital headache and hypoesthesia in the V1 region.

Subbasal corneal nerve damage in patients with bacterial keratitis: in vivo confocal microscopy study.

PURPOSE: To examine subbasal corneal nerve changes in patients with bacterial infectious keratitis using in vivo confocal microscopy. METHODS: Thirteen patients (13 eyes) with unilateral bacterial keratitis and 12 healthy controls were prospectively enrolled in the study. In vivo confocal microscopy was performed in all the patients at 2 time points, in the acute phase of infectious keratitis and at 28 ± 0.6 months after resolution of the infection. RESULTS: The subbasal nerve length was 5.15 ± 1.03 mm/mm2 during the acute phase of bacterial keratitis (compared with that of the controls: 19.02 ± 1.78 mm/mm2, p<0.05). Despite the significant corneal nerve regeneration over the interval of 28 months after the resolution of the infection, the nerve density was still significantly reduced as compared with that of the controls (9.73 ± 0.93 mm/mm2, p<0.05). Moreover, in vivo confocal microscopy images showed diffuse high-reflecting areas referring to the scar tissue areas with thin and tortuous nerve branches regenerating toward these areas. CONCLUSIONS: A partial corneal nerve regeneration of subbasal nerve plexus during the first 28 months after the acute phase of infectious keratitis was observed. Moreover, the regenerated nerves of the patients remained morphologically altered as compared with those of the healthy controls. These results may be relevant to the clinical follow-up and surgical planning for these patients.

Optogenetic Inhibition of Nav1.8 Expressing Corneal Afferents Reduces Persistent Dry Eye Pain.

Purpose: The aim of the present study was to investigate the contribution of Nav1.8 expressing corneal afferent neurons to the presence of ongoing pain in lacrimal gland excision (LGE)-induced dry eye. Methods: The proton pump archaerhodopsin-3/eGFP (ArchT/eGFP) was conditionally expressed in corneal afferents using Nav1.8-cre mice. Dry eye was produced by unilateral LGE. Real time place preference was assessed using a three-chamber apparatus. A neutral, unlit center chamber was flanked by one illuminated with a control light and one illuminated with an ArchT activating light. For real-time preference, animals were placed in the neutral chamber and tracked over five 10-minute sessions, with the lights turned on during the second and fourth sessions. In other studies, movement was tracked over three 10-minute sessions (the lights turned on only during the second session), with animals tested once per day over the course of 4 days. A local anesthetic was used to examine the role of ongoing corneal afferent activity in producing place preference. Results: The corneal afferent nerves and trigeminal ganglion cell bodies showed a robust eGFP signal in Nav1.8-cre;ArchT/eGFP mice. After LGE, Nav1.8-cre;ArchT/eGFP mice demonstrated a preference for the ArchT activating light paired chamber. Preference was prevented with pre-application to the cornea of a local anesthetic. Nav1.8-cre;ArchT/eGFP mice with sham surgery and LGE wild-type control mice did not develop preference. Conclusions: Results indicate LGE-induced persistent, ongoing pain, driven by Nav1.8 expressing corneal afferents. Inhibition of these neurons represents a potential strategy for treating ongoing dry eye-induced pain.

A whole-mount immunohistochemistry protocol for detection of mouse corneal nerves.

A cornea is innervated by sensory nerves, which branch into thick trunks, subbasal plexuses, and sensory endings. Appropriate assessment of nerve structure in a tissue provides a more complete understanding of the role of nerves in health and disease. Here, we present a whole-mount immunohistochemistry protocol that facilitates evaluation of nerve architecture throughout the mouse cornea. The fixation step in this protocol allows for reliable detection of nerve structures within the cornea and likely other tissues. For complete details on the use and execution of this protocol, please refer to Yun et al, (2020).

The pattern of the inferocentral whorl region of the corneal subbasal nerve plexus is altered with age.

PURPOSE: To describe the pattern of the nerves in the inferocentral whorl region of the human corneal subbasal nerve plexus (SBNP) in health and diseases known to affect the subbasal nerves. METHODS: Laser-scanning in vivo confocal microscopy (IVCM) was used to image the SBNP bilaterally in 91 healthy subjects, 39 subjects with type 2 diabetes mellitus (T2DM), and 43 subjects with Parkinson's disease (PD). Whorl regions were classified according to nerve orientation relative to age and health/disease status. RESULTS: Of 346 examined eyes, 300 (86.7%) had an identifiable whorl pattern. In healthy subjects, a clockwise nerve orientation of the whorl was most common (67.9%), followed by non-rotatory or 'seam' morphology (21.4%), and counterclockwise (10.7%). The clockwise orientation was more prevalent in healthy subjects than in T2DM or PD (P < 0.001). Healthy individuals below 50 years of age had a predominantly clockwise orientation (93.8%) which was reduced to 51.9% in those over 50 years (P < 0.001). Age but not disease status explained whorl orientation in T2DM and PD groups. Moreover, whorl orientation is bilaterally clockwise in the young, but adopts other orientations and becomes asymmetric across eyes with age. Finally, we report reflective 'dot-like' features confined to the whorl region of the subbasal plexus, sometimes appearing in close association with subbasal nerves and present in 84-93% of examined eyes regardless of disease status, eye or sex. CONCLUSION: Subbasal nerves in the inferocentral whorl region are predominantly clockwise in young, healthy corneas. With aging and conditions of T2DM and PD, counterclockwise and non-rotatory configurations increase in prevalence, and bilateral symmetry is lost. Mechanisms regulating these changes warrant further investigation.