Experimental Strategies

Optogenetics

DREADDs

Immunohistochemistry

Behavioral assays

Calcium imaging in living brain slices

Confocal & multiphoton microscopy

Brain slice electrophysiology

Two stereotaxic surgical stations with dissecting optics and virus delivery systems. These surgical stations are equipped with rat/mouse digital stereotaxic frames (Stoelting; Kopf), micromanipulators, isoflurane vaporizers and injectors (Drummond Nanoinjectors and MPI Sutter) necessary for intracranial transfection of virus and/or fluorescent microspheres. An adjacent bench is used for IV-catheter surgeries for self-administration experiments. 

Sixteen operant chambers outfitted for drug and sucrose self-administration studies.  Chambers are used for behavioral testing of transgenic and wild type rodents. In addition to self-administration studies, these chambers support progressive ratio work. We are outfitted for behavioral testing (von Frey) in spared-nerve injury (SNI) and inflammatory pain models. We are working closely with the Comprehensive Pain and Addiction Center (CPAC), which offers additional testing equiptment for CPP and locomotor activity equipped with Ethovision (Noldus Technologies) for real-time tracking of behavior. 

Calcium imaging in living brain slices. Commercially built two-photon Bruker Ultima live imaging station equipped with a coherent laser on a anti-vibration (9’x5’) optical table (Newport). This imaging system is integrated with slice electrophysiology rig and incorporates conventional galvanometer-based scanners, standard imaging set and uncaging-optics. The imaging system is integrated with upright microscope (Olympus) equipped with infra-red illumination and a video camera. A custom stage holding a brain slice chamber and 2 micromanipulators (Sikiyou). The microscope has a 5X, 40X (water-immersion) and 60X (water-immersion) objective lens. We are working closely with Dr. Khanh Kieu (UA optical Sciences) to optimize and integrate light sheet microscopy approaches using low power lasers for imaging neural circuitry relevant to motivated behaviors.

Histology and fluorescent immunohistochemistry station. Space includes a Leica microtome, stereomicroscope with camera, a purpose-built gene gun, shakers, tissue processing stations, and water baths and slide warmer fully sufficient to support the fluorescent immunohistochemistry. 

Confocal microscopy. Our Department possess two confocal microscopes for analysis of brain circuits and quantitative immunohistochemistochemical studies in fixed tissue. The microscopes include an Olympus Fluoview FV1200 equipped with both HeNe and Ar lasers with multiple excitation and emission fluorescence filters and Fluoview software. Also available is a Zeiss LSM 700 AXIO Observer.Z1 Confocal Microscope system, with Zeiss ZEN Imaging software. Both systems are networked to Dell Precision workstations.

Two brain slice electrophysiology rigs. These standalone rigs are outfitted with an Olympus BX51 microscopes with optics for differential interference contrast (DIC) and fluorescent imaging. Rigs possess amplifiers (Axopatch 700B) for patch-clamp recordings in brain slices, iontophoresis units and LEDs for optogenetic photostimulation (i.e., ChR2) or UV uncaging of compounds including caged-calcium.