Envisu r4310

We performed OCT (Envisu R4310, Leica, Wetzlar, Germany) as previously described (Miwa et al., 2019a (link)). Briefly, a combination of 0.5% tropicamide and 0.5% phenylephrine (Santen Pharmaceutical, Osaka, Japan) was added to mice for 5 min for induction of mydriasis. Anesthetized mice by a combination of MMB were placed in an OCT platform and then OCT was immediately conducted. 0.2, 0.4 and 0.6 mm from the optic nerve head in the retina were marked. Then, total retinal thickness (from the outer retina to the inner retina including the ganglion cell layer) was measured via OCT software (Envisu R4310, Leica, Wetzlar, Germany).

The efraction and the ocular components were measured using an infrared photorefractor (Steinbeis Transfer Center, Tübingen, Germany) and an SD-OCT system (Envisu R4310, Leica, Wetzlar, Germany). All measurements were performed under the condition of mydriasis by 0.5% tropicamide and 0.5% phenylephrine combination eye drops (Santen, Osaka, Japan), and general anesthesia with the combination of midazolam (Sandoz K.K., Tokyo, Japan), medetomidine (Domitor^®, Orion Corporation, Espoo, Finland) and butorphanol tartrate (Meiji Seika Pharma Co., Ltd., Tokyo, Japan) (MMB). A 0.01 mL/g dose of MMB was administered intraperitoneally. The refractive error values were averaged over 100 measurements. The AL was determined from the anterior corneal surface to the retinal pigment epithelium along the corneal vertex reflection. The CCT was the distance from the anterior corneal surface to the posterior corneal surface, the ACD was from the posterior corneal surface to the anterior lens surface, and the LT was from the anterior lens surface to the posterior lens surface. The CCR was measured by an infrared keratometer (Steinbeis Transfer Center, Tübingen, Germany) [50 (link)]. The lens elasticity was measured by Softmeasure (Horiuchi Electronics Co. LTD., Tokyo, Japan) after enucleation under an overdose of anesthesia.

AL was measured from the anterior corneal surface to the retinal pigment epithelium using a spectral domain optical coherence tomography (SD-OCT) system (Envisu R4310, Leica, Germany). The refraction was detected using an eccentric infrared photo refractor (Steinbeis Transfer Center, Germany) at the vertical pupil meridian. The data were automatically recorded by the program when the parameter values were stable. Choroidal thickness (ChT) was measured using SD-OCT, and the posterior surface of the choroid was quantified using Image J (Ver 1.53, NIH). ChT was determined using the formula: area divided by circumference. Intraocular pressure (IOP) was measured using a tonometer (Tono Lab, Icare Finland Oy, Vantaa, Finland) calibrated for mice under anaesthesia. Parts images of ocular components measurement were shown in Additional file 5: Fig. S5.
Mice were anesthetized with 0.75 mg/kg medetomidine (Sandoz K.K., Tokyo, Japan), 4 mg/kg midazolam (Domitor®, Orion Corporation, Espoo, Finland), and 5 mg/kg butorphanol tartrate (Meiji Seika Pharma Co., Ltd., Tokyo, Japan) dissolved in normal saline. Mydriasis was induced by 0.5% tropicamide eye drops (Santen, Osaka, Japan), and mice were placed in a cylindrical holder for measurement.