Search

Background: Molecular autopsy in sudden unexplained death in the young (SUDY) victims cannot usually be performed if tissue suitable for DNA extraction is not retained at autopsy. Objective: The purpose of this study was to assess the feasibility and clinical value of posthumous genetic testing for long QT syndrome (LQTS) using residual material from the neonatal screening (Guthrie) card in SUDY victims. Methods: Twenty-one cases were investigated up to 13 years after death. Deaths occurred at <1 year in one, 1–18 years in 18, and 19–35 years in two patients. Guthrie cards were 3–39 years old. DNA was extracted, and amplicons corresponding to the coding regions of the LQTS genes 1, 2, 3, 5, and 6 underwent either denaturing high-performance liquid chromatography screening or direct DNA sequencing. Results: Adequate DNA was extracted in every case, although repeated purification and amplification was often required. Rare variants were detected in six of 19 cases undergoing diagnostic screening. Four (21%) are considered to be pathological and have been used for family screening: R243C and H455Y in KCNQ1 in 12-year-old and 13-year-old boys, respectively, and Q81H and S621R in KCNH2 in 21-month and 28-year-old females, respectively. Variants of uncertain significance were R1047L in KCNH2 in a 2-year-old girl and S38G in KCNE1 in a 19-month-old boy. Point mutation tests for previously identified familial LQTS mutations revealed a positive result in both cases: E146K in KCNQ1 and exon 6-4del in KCNH2. Conclusion: Residual material from Guthrie cards collected for newborn metabolic screening can be used as a reliable source of DNA for the posthumous diagnosis of LQTS decades after SUDY, although purification and amplification of DNA is time intensive. [Copyright &y& Elsevier]

Immunoreactivity of GABAA receptor subunits and the receptor anchoring protein gephyrin was investigated in the human globus pallidus using antibodies raised against the α1 and γ2 subunits of the GABAA receptor complex and gephyrin. The results revealed increased GABAA receptor subunit immunoreactivity and unchanged levels of gephyrin immunoreactivity in Huntington''s diseased (HD) globus pallidus (GP). The results demonstrate that gephyrin immunoreactivity did not change in unison with GABAA receptor changes in HD, suggesting that the receptor anchoring protein gephyrin is unaltered and maintains a stable lattice structure in the face of GABAA receptor changes in HD. [Copyright &y& Elsevier]

Gephyrin is an ubiquitously expressed protein that, in the central nervous system, generates a protein scaffold to anchor inhibitory neurotransmitter receptors in the postsynaptic membrane. It was first identified as a protein component of the glycine receptor complex. Recent studies have demonstrated that gephyrin is colocalized with several subtypes of GABAA receptors and is part of postsynaptic GABAA receptor clusters. Here, we describe a study of the regional and cellular distribution of gephyrin in the human brain, determined by immunohistochemical localisation at the light and confocal laser scanning microscopic levels. At the regional level, gephyrin immunoreactivity was observed in most of the major brain regions examined. The most intense staining was in the cerebral cortex, hippocampus and caudate-putamen, in various brainstem nuclei with more moderate levels in the thalamus and cerebellum. At the cellular level gephyrin immunoreactivity was present on the plasma membranes of the soma and dendrites of pyramidal neurons throughout the various cortical regions examined. In the hippocampus, intense staining was observed on the granule cells of the dentate gyrus, and neurons of the CA1 and CA3 regions showed intense punctate gephyrin staining on their apical dendrites and cell bodies. Gephyrin immunoreactivity was also observed on neurons in the thalamus, globus pallidus and substantia nigra. In the putamen intense labelling of the striosomes was observed; most of the medium-sized neurons in the caudate-putamen were weakly labelled and many large neurons of the striatum were conspicuously stained. Many of the brainstem nuclei, notably the dorsal motor nucleus of the vagus, hypoglossal nucleus, trigeminal nucleus and inferior olive were all labelled with gephyrin. The spinal cord also showed high levels of gephyrin immunoreactivity. Our results demonstrate that the anchoring protein gephyrin is ubiquitously present in the human brain. We therefore suggest that gephyrin may have a central organizer role in assembling and stabilizing inhibitory postsynaptic membranes in human brain and is similar in function to those observed in the rodent brain. These findings contribute towards elucidating the role of gephyrin in the human brain. [Copyright &y& Elsevier]