Stereo lumar v12 fluorescence microscope

Xenopus laevis eggs were collected and fertilized in vitro, as previously described^{9 (link)}. A total of 100 pg of DNA, consisting of 50 pg of pcDNA4TO-eGFP, pcDNA4TO-eG-SCON-FP or pcDNA4TO-eG-SC-FP, and 50 pg of an mCherry-expressing plasmid, was injected into two dorso-animal blastomeres of X. laevis embryos at the 4-cell stage, and imaging was performed approximately 24 h later at the early tailbud stage. For Danio rerio experiments, 12 pg of DNA in total, consisting of 6 pg of pcDNA4TO-eGFP, pcDNA4TO-eG-SCON-FP or pcDNA4TO-eG-SC-FP, and 6 pg of mCherry-expressing plasmid, was injected into each fertilized egg/1-cell stage D. rerio embryo, and imaging was performed approximately 24 h later. X. laevis embryos were imaged using a stereomicroscope (Leica, M165MC) equipped with GFP (excitation: ET470/40 nm; emission: ET525/50 nm) and mCherry (excitation: ET560/40 nm; emission: ET630/75 nm) filters. D. rerio embryos were imaged with a Stereo Lumar.V12 fluorescence microscope (Zeiss) equipped with GFP (excitation 470/40 nm; emission 525/50 nm) and mCherry (excitation 550/25 nm; emission 605/70 nm) filters.

Surgical procedures in male Sprague–Dawley rats (283.25 ± 48.42 g body weight, mean ± standard deviation) were performed as reported [36 (link)]. Rats were deeply anesthetized by intraperitoneal administration of ketamine (100 mg/mL, 0.08 mL/100 g) and xylazine (20 mg/mL, 0.06 mL/100 g). The tail vein was catheterized, and animals were placed in a stereotactic frame under a SteREO Lumar V12 fluorescence microscope (Zeiss Ltd., Oberkochen, Germany). Body temperature was continuously monitored and kept stable at 37 ± 0.5 °C using a feedback-controlled heating pad (Physitemp Ltd., Clifton, NJ USA). Heart rate, breath rate, and oxygen saturation levels were continuously monitored using MouseOx (STARR Life Sciences Ltd. Oakmont, PA, USA). A cranial bone section (4 mm caudal, 2 mm frontal, 5 mm lateral to bregma) was opened over the right sensory-motor cortex. The dura and arachnoid layers were removed, and the exposed cortex was continuously perfused with artificial cerebrospinal fluid (ACSF) containing (in mM): 124 NaCl, 26 NaHCO₃, 1.25 NaH₂PO₄, 2 MgSO₄, 2 CaCl₂, 3 KCl, and 10 Glucose (pH 7.4).