30, Schneeberger) and

driven by pneumatic linear actuators (6604k11, McMaster Carr). A custom implantable titanium headplate was designed to mate with the kinematic clamp. Low-force, miniature snap action switches (D42L-R1XL, Cherry) mounted on the clamp were used to detect when the anterior edge of the headplate reached the rear end of the heaplate slot. When the headplate was in this location, actuation of the kinematic clamp would drive the piston-mounted ball bearings toward the conical depression and the V groove. ZD1839 clinical trial Interaction between the piston-mounted ball bearings and the conical depression and the V groove constrained 5 of 6 of the degrees of freedom of the headplate. The sixth degree, pitch, was constrained by interactions between the top plane of the headplate and the flat surface of the ceiling of the headplate slot. In some cases, we found it necessary to more firmly constrain pitch as some animals were able to produce sufficient torque on their headplate to create pitch movements, which contributed to brain motion. In these cases, we mounted miniature Teflon-tipped brass arms to the pistons that contacted the underside of the anterior portion of the headplate and clamped the headplate more firmly to the ceiling of the headplate slot. Delivery of air pressure (0–90 PSI) to the pneumatic linear actuators was controlled either by a manual regulator combined with two solenoid valves (T9-65-900, Toohey) or using

a voltage-controlled regulator (ITV1050-31N2S4, SMC). The kinematic clamp was installed along one wall of the modified rat BKM120 operant conditioning chamber (Island Motion), which also contained two additional reward pokes on the left and right side. Position of the center poke was controlled either by a custom manual translation stage or using a motorized linear stage (ET-50-21, Newmark Systems). A behavioral control system, Bcontrol (see below), controlled the timing of fluid reward and auditory and

visual cues. A schematic diagram of the overall system architecture is shown in Figure S1. The operant conditioning chamber was mounted on an air table that also housed a movable objective microscope (Sutter Instruments). The microscope was positioned so tuclazepam that the vertically oriented objective was centered over the headport clamp. Two objectives were used for TPM: a 40×, 0.8 NA, water immersion (LUMPLFN40XW, Olympus) and a 40×, 0.6 NA with a correction collar (LUCPLFLN40X, Olympus). A Ti:Sapphire laser was used as an illumination source (Chameleon Ultra II, Coherent) for 920 nm light. A delrin collar was designed to mount on the barrel of the water-immersion objective and position two stainless steel tubes, one for fluid delivery and one for fluid removal. Upon activation of the clamp at the beginning of each insertion event, 75 μl of immersion fluid was delivered to the gap between the implanted optical window and the face of the imaging objective. Inflow was produced by a gravity-fed system.