Judith H. Ford

The human glycine receptor β subunit : primary structure, functional characterisation and chromosomal localisation of the human and murine genes

The inhibitory glycine receptor (GlyR) is a pentameric receptor comprised of alpha and beta subunits, of which the beta subunit has not been characterised in humans. A 2106 bp cDNA, isolated from a human hippocampal cDNA library, contained an open reading frame of 497 amino acids which encodes the beta subunit of the human GlyR. The mature human GlyR beta polypeptide displays 99% amino acid identity with the rat GlyR beta subunit and 48% identity with the human GlyR alpha 1 subunit. Neither [3H]strychnine binding nor glycine-gated currents were detected when the human GlyR beta subunit cDNA was expressed in the human embryonic kidney 293 cell line. However, co-expression of the beta subunit cDNA with the alpha 1 subunit cDNA resulted in expression of functional GlyRs which showed a 4-fold reduction in the EC50 values when compared to alpha 1 homomeric GlyRs. Glycine-gated currents of alpha 1/beta GlyRs were 17-fold less sensitive than homomeric alpha 1 GlyRs to the antagonists picrotoxin, picrotoxinin and picrotin, providing clear evidence that heteromeric alpha 1/beta GlyRs were expressed. The beta subunit appears to play a structural rather than ligand binding role in GlyR function. Fluorescence in situ hybridisation was used to localise the gene encoding the human GlyR beta subunit (GLRB) to chromosome 4q32, a position syntenic with mouse chromosome 3. In situ hybridisation using the human GlyR beta subunit cDNA showed that the murine GlyR beta subunit gene (Glrb) maps to the spastic (spa) locus on mouse chromosome 3 at bands E3-F1. This is consistent with the recent finding that a mutation in the murine GlyR beta subunit causes the spa phenotype. It also raises the possibility that mutations in the human beta subunit gene may cause inherited disorders of the startle response.

A fluorescent in situ hybridization analysis of the chromosome constitution of ejaculated sperm in a 47, XYY male

Two semen samples from a 47, XXY male were examined using chromosome‐specific DNA probes and fluorescent in situ hybridization (FISH) to determine the distribution of sex chromosomes and an autosome (chromosome 17) in the sperm. A motile population of sperm was also prepared from one sample using the swim‐up technique to compare the motile and total sperm populations. Chromosomes were localized using single FISH and a biotinylated chromosome 17 probe (TR17), or double FISH using a biotinylated X chromosome probe (TRX) and a digoxigenin‐labelled Y chromosome probe (HRY). Labelling efficiencies were 95–98%. Ploidy levels were estimated by measurement against a microscope eyepiece graticule. The overall ratio of X‐to Y‐bearing sperm was 47% to 48.4% in the neat samples, and 48.4% to 45.3% in the swim‐up fraction. Neither of the ratios was significantly different from 1:1. The frequencies of monosomic and disomic (but otherwise haploid sperm) were not different from the frequencies we observed in normal donors. In contrast, the frequencies of both diploid and tetraploid cells were increased in the neat samples of the XYY male. In the swim‐up fractions, however, none of these parameters differed from those of ten normal semen donors. These results support the hypothesis that the extra Y chromosome in XYY men is eliminated during spermatogenesis.

Detection of X- and Y-bearing human spermatozoa after motile sperm isolation by swim-up

OBJECTIVE To assess the ratio of X- to Y-bearing human spermatozoa in motile fractions isolated by the swim-up technique. DESIGN The proportions of X- and Y-bearing sperm were determined in neat semen samples (control) and in motile fractions isolated from the same samples by swim-up. X- and Y-bearing sperm were simultaneously identified using chromosome-specific DNA probes and double fluorescence in situ hybridization. SETTING Hospital-based university department. PARTICIPANTS Ten healthy donors with normal semen characteristics. MAIN OUTCOME MEASURES The distribution of haploid cells (X or Y), normal size cells with two sex chromosome (XX, YY, or XY), and large cells containing two (XX, YY, or XY) or four (XXYY) sex chromosomes were measured in neat semen samples and in motile fractions prepared by swim-up. RESULTS Overall, 95% of sperm in the neat semen and swim-up fractions were labeled with the probes. The ratios of X- to Y-bearing sperm were 47.3:46.9 (neat semen) and 48.4:47.1 (swim-up fractions), which were not significantly different from a 1:1 ratio. The frequencies of sperm with normal size nuclei and two sex chromosomes (XX, YY, or XY) in the swim-up fractions were not significantly different from the controls, but there was a significant reduction in the proportion of cells with large nuclei and two (XX, YY, or XY) or four (XXYY) sex chromosomes in the swim-up fractions. CONCLUSIONS The swim-up technique does not selectively enrich either X- or Y-bearing sperm. Because the isolation of motile spermatozoa is an important procedure for routine IUI, IVF-ET, and GIFT, the results of this study are important reassurance that the sex ratio is not altered by this method of sperm preparation.

Screening of karyotype and semen quality in an artificial insemination program: acceptance and rejection criteria.

Semen quality and karyotype were screened in all men offering to be donors for an artificial insemination (AID) program. The criteria for accepting or rejecting semen have now been set with respect to this sample of the population. There was no evidence of differences between the pregnancy rates of the accepted donors. One of 172 potential donors with a clear medical history had a chromosomal abnormality, 4 had pericentric inversions of chromosome 9, and 14 had other heterochromatic variants. Of the recipients of AID, 5 of 196 women had chromosomal abnormalities, and 12 had heterochromatic variants.

Unusual segregation in a family with a 11/21 translocation

A familial 11/21 translocation is described where the proband has an unbalanced translocation and the oldest translocation carrier shows mosaicism with a partial trisomy no. 21.

Aneuploidy Studies in Sperm: Post-Meiotic Selection against Aneuploid Sperm

Observations on the parental transmission of chromosomes in trisomic conceptions have shown that the disomic contribution is usually maternal in origin (Hassold & Takaesu, 1989). It is well known that trisomic conceptions increase with maternal age and it is usually assumed that this reflects an increase in meiotic error with maternal age. No equivalent age-related change in aneuploidy with paternal age is known. If there is no such change, then the proportion of male contributions to trisomy should decrease with increasing maternal age (Sved & Sandler, 1981) unless the rate of trisomie conceptions is the same at all ages and the maternal age effect is post-meiotic, possibly one of decreased early recognition or correction of trisomic conceptions.