ABSTRACT
There is a growing effort to image single neurons in vivo, and observe their individual contribution to the brain's functional organization. This effort generally relies on two-photon imaging to explore the structure and activity of cortical columns extending beneath the brain's surface. The need to protect living tissue, however, demands the introduction of coverslips and similar objects that can modify the optics of the imaging beam. This paper develops three-dimensional (3D) analytical and numerical models to characterize and correct for the resulting degradation of image quality. We have illustrated the use of these models by describing a simple, practical technique to reduce the effect of spherical aberration for in vivo two-photon fluorescence experiments.
Subject(s)
Microscopy, Fluorescence, Multiphoton/methods , Animals , Brain/cytology , Brain/physiology , Computer Simulation , Functional Neuroimaging/methods , Functional Neuroimaging/statistics & numerical data , Imaging, Three-Dimensional/methods , Imaging, Three-Dimensional/statistics & numerical data , Mathematical Concepts , Microscopy, Fluorescence, Multiphoton/statistics & numerical data , Neurons/cytology , Neurons/physiology , Optical PhenomenaABSTRACT
PURPOSE: Paragangliomas of the head and neck are neuroendocrine tumors and are associated with germ line mutations of the tricarboxylic acid cycle-related genes SDHB, SDHC, SDHD, and SDHAF2. Hypoxia is important in most solid tumors, and was directly implicated in tumorigenesis over 40 years ago when it was shown that dwelling at high altitudes increases the incidence of carotid body hyperplasia and paragangliomas. Although recent research has now elucidated several pathways of hypoxia in paragangliomas, nothing is currently known of the genetics or of gene-environment interactions in high-altitude paraganglioma. We postulated that SDH mutations might play a role in these tumors. EXPERIMENTAL DESIGN: Patients from a Mexican family, originating and resident in Guadalajara, were tested for mutations of SDHD, and subsequently, for mutations of SDHB followed by immunohistochemical confirmation of SDHB loss. RESULTS: Two patients, born and resident at altitudes of between 1,560 and 2,240 m, were found to have head and neck paragangliomas, including a remarkably aggressive recurrent tumor. Mutation analysis identified a pathogenic missense mutation in exon 7 of SDHB, c.689G>A, p.Arg230His, and loss of the SDHB protein was confirmed by immunohistochemistry. CONCLUSIONS: This is the first report of a SDH gene mutation in paraganglioma at high altitude. A rapidly recurrent head and neck paraganglioma is a very rare finding in an SDH mutation carrier, suggesting a gene-environment interaction. Neither patient showed evidence of sympathetic paraganglioma.