Abstracts

Hyperekplexia (HEK) is a rare neurogenetic disorder characterized by an exaggerated startle response. HEK, predominantly an autosomal dominant disease, although recessive and sporadic cases are known, is due to mutations in the gene (5q32) encoding the alpha subunits of the glycine receptor (GLRA1) that facilitates inhibitory neurotransmission.
A consanguineous family with HEK from Sultan Qaboos Medical College, Muscat, Oman underwent clinical, DNA and functional studies.(NH) Out of 7 children 4 males had the major form of HEK. The index case is a 14-year-old male diagnosed with [ldquo]seizures[rdquo] on the second post-natal day with startle response, spasms in response to crying, and hypertonia. Laboratories studies were: normal chemistries, lactate, ammonia, urine, plasma amino acids, and TORCH. EEG showed spasms not associated with spike-wave discharges. MRI/CT scan of the brain was normal. He was treated with Clonazepam with improvement. Three brothers, ages 9 and 5 years, and 2 months are also affected. EEG studies show no inter-ictal transients but slow background. DNA was isolated from peripheral blood lymphocytes. The coding region of both GlyR [beta] gene (GLRB) and GlyR [alpha]-1 gene were sequenced 1 kb upstream and downstream from each 5[apos] and 3[apos] end of this gene. The exons were amplified by 16 sets of primers designed to cover the entire coding region and 1 kb upstream and downstream of the gene. Amplicon amplification and sequence reaction were conducted following a standard protocol [Butler]. Dideoxy sequencing of the forward and reverse DNA strand of each individual was performed using a 3100 sequencer (Applied Biosystems, USA).
Evaluation of functional binding of the variance found at CREB site in the glycine receptor gene alpha was carried out using the electrophoric mobility shift assay (EMSA] (Active motif) [italic]DNA studies:[/italic] Sequencing of the promoter and coding regions of GLRA1 revealed two single nucleotide sequence substitutions . A novel 420C[gt]T substitution was detected in the promoter region upstream of exon-1 and the common nucleotide polymorphism -357C[gt]T (gi:51511721) was also observed. All 3 affected were homozygous for the 113T [amp] 176T alleles and both unaffected parents were heterozygous.
Analysis of nuclear factor binding CREB promoter sequence at position 176 was labeled on the coding strand at the 5[apos] end, labeled with the DNA probe (1.5 [times] 104 cpm) incubated with nuclear extract derived from HEK cells. A supershift was seen at the lane where probe plus antibody and nuclear extract were mixed. This is the first report of an Arabic family with autosomal recessive major form of HEK with novel sequence variant in GLRA1 promoter region.
The variant found at -176 at the promoter region of the glycine receptor is a CREB site and it may be involved in the loss of function seen in HEK. (Supported by Epilepsy Foundation of America Holden Research Award-- Miguel E. Fiol M.D.)