The Optical Fractionator Technique to Estimate Cell Numbers in a Rat Model of Electroconvulsive Therapy

TY - JOUR

T1 - The Optical Fractionator Technique to Estimate Cell Numbers in a Rat Model of Electroconvulsive Therapy

AU - Olesen, Mikkel Vestergaard

AU - Needham, Esther Kjær

AU - Pakkenberg, Bente

PY - 2017/7/9

Y1 - 2017/7/9

N2 - Stereological methods are designed to describe quantitative parameters without making assumptions about size, shape, orientation and distribution of cells or structures. These methods have been revolutionary for quantitative analysis of the mammalian brain, in which volumetric cell populations are too high to count manually, and stereology is now the technique of choice whenever estimates of three-dimensional quantities need to be extracted from measurements on two-dimensional sections. All stereological methods are in principle unbiased; however, they rely on proper knowledge about the structure of interest and the characteristics of the tissue. Stereology is based on Systematic Uniformly Random Sampling (SURS), with adjustment of sampling to the most efficient level in respect to precision, providing reliable, quantitative information about the whole structure of interest. Here we present the optical fractionator in conjunction with BrdU immunohistochemistry to estimate the production and survival of newly-formed neurons in the granule cell layer (including the sub-granular zone) of the rat hippocampus following electroconvulsive stimulation, which is among the most potent stimulators of neurogenesis. The optical fractionator technique is designed to provide estimates of the total number of cells from thick sections sampled from the full structure. Thick sections provide the opportunity to observe cells in their full 3-D extent and thus, allow for easy and robust cell classification based on morphological criteria. When correctly implemented, the sensitivity and efficiency of the optical fractionator provides accurate estimates with a fixed and predetermined precision.

AB - Stereological methods are designed to describe quantitative parameters without making assumptions about size, shape, orientation and distribution of cells or structures. These methods have been revolutionary for quantitative analysis of the mammalian brain, in which volumetric cell populations are too high to count manually, and stereology is now the technique of choice whenever estimates of three-dimensional quantities need to be extracted from measurements on two-dimensional sections. All stereological methods are in principle unbiased; however, they rely on proper knowledge about the structure of interest and the characteristics of the tissue. Stereology is based on Systematic Uniformly Random Sampling (SURS), with adjustment of sampling to the most efficient level in respect to precision, providing reliable, quantitative information about the whole structure of interest. Here we present the optical fractionator in conjunction with BrdU immunohistochemistry to estimate the production and survival of newly-formed neurons in the granule cell layer (including the sub-granular zone) of the rat hippocampus following electroconvulsive stimulation, which is among the most potent stimulators of neurogenesis. The optical fractionator technique is designed to provide estimates of the total number of cells from thick sections sampled from the full structure. Thick sections provide the opportunity to observe cells in their full 3-D extent and thus, allow for easy and robust cell classification based on morphological criteria. When correctly implemented, the sensitivity and efficiency of the optical fractionator provides accurate estimates with a fixed and predetermined precision.

KW - Animals

KW - Cell Count/methods

KW - Disease Models, Animal

KW - Electroconvulsive Therapy/methods

KW - Immunohistochemistry/methods

KW - Male

KW - Rats

U2 - 10.3791/55737

DO - 10.3791/55737

M3 - Journal article

C2 - 28715378

SN - 1940-087X

VL - 125

JO - Journal of Visualized Experiments

JF - Journal of Visualized Experiments

M1 - e55737

ER -