Using an approach that minimizes sampling bias of spines, dendritic segments (20 - 25 μm in length) were selected with a systematic-random design (Duan et al., 2003; Hao et al., 2006; Radley et al., 2006) and imaged essentially as described on a Zeiss LSM 510 confocal microscope (Zeiss, Thornwood, NY, USA) using a 100X/1.4 N.A. Plan-Apochromat objective with a digital zoom of 3.5 and an Ar/Kr laser at an excitation wavelength of 488 nm. All confocal stacks were acquired at 512 × 512 pixel resolution with a z-step of 0.1 μm. All confocal stacks included approximately 1 μm above and below the identified dendritic segment. On average, three z-stacks were imaged per neuron. In order for a dendritic segment to be optically imaged, it had to satisfy the following criteria: (1) the entire segment had to fall within a depth of 50 μm; (2) dendritic segments had to be either parallel or at acute angles to the coronal surface of the section; and (3) segments did not overlap other segments that would obscure visualization of spines (Radley et al., 2006;
