Molecular forms of gonadotropin-releasing hormone associated peptide (GAP): changes within the rat hypothalamus and release from hypothalamic cells in vitro

We have developed RIAs using antisera directed against the cryptic peptide of the GnRH precursor (termed GnRH-associated peptide, GAP) and have used these together with a GnRH assay to characterize proGnRH-derived peptides in rat hypothalamic extracts. On Sephadex chromatography we have identified three molecular forms of GAP-like immunoreactivity (GAP-LI), in addition to the GnRH decapeptide. The largest of these forms is an 8.0-kilodalton (kD) GAP-LI which appears to be the complete proGnRH peptide. The second is a 6.5-kD GAP-LI, and is similar to the complete cryptic peptide (i.e. proGnRH14-69 or GAP1.56). The third peptide is a 2.5 kD C-terminal fragment of the cryptic peptide, representing a processed form of GAP. In whole hypothalamic extracts from normal rats the 8.0-kD form was the major form, comprising 60-70% of the total GAP-LI. All three forms were present in three distinct areas of the rat hypothalamus, namely median eminence (ME), anterior and mid-hypothalamus. However in the ME the proportion of 8.0-kD GAP-LI was significantly reduced and the proportion of 6.5-kD GAP-LI significantly increased compared to anterior and mid-hypothalamic samples (p less than 0.05). In whole hypothalamic extracts from pregnant and lactating rats the total content of proGnRH-derived peptides was reduced but the relative proportions of these peptides were not significantly changed from normal female rats. However, in postlactating rats, 2 weeks after removal of pups, the total levels of GAP-LI were unchanged compared to normals, but the percentage of 8.0-kD GAP-LI was significantly decreased and the percentage of 2.5-kD GAP-LI significantly increased compared to normals (p less than 0.05), suggesting that proGnRH may undergo additional processing dependent on physiological condition. In fetal and neonatal rats the proportion of the 6.5-kD peptide was increased and that of the 8.0-kD peptide decreased compared to adults, and this change became less significant with increasing age. In ovariectomized rats the proportion of 6.5-kD GAP-LI was increased and that of 8.0-kD GAP-LI decreased; this change was partially reversed with steroid treatment. Both the 6.5 and 2.5-kD forms were released by high K+ stimulation of neonatal hypothalamic cells in culture. These results indicate that there is differential processing of the proGnRH precursor within the hypothalamus and in altered physiological states.     

Post-translational processing of thyrotropin-releasing hormone precursor in rat brain: identification of 3 novel peptides derived from pro TRH

The sequence of rat hypothalamic pro-thyrotropin releasing hormone, deduced by sequencing of cDNA, in addition to 5 TRH progenitor genitor sequences contains leader, trailer and 4 intervening sequences separated by paired basic amino acid sequences. We have developed radioimmunoassays to synthetic peptides corresponding to portions of these cryptic proTRH sequences and have used these assays to identify and partially characterize proTRH peptides, distinct from TRH, in extracts of rat brain. Two of these peptides correspond closely in size to one intervening sequence and the car☐y-terminal sequence of proTRH. Three other peptides correspond to the intact amino-terminal leader sequence and two peptides formed by a further cleavage of the leader sequence at an internal paired basic amino acid sequence.     

A new feature of the ablepharon macrostomia syndrome: zygomatic arch absence

The ablepharon macrostomia syndrome is an extremely rare congenital anomaly. It is characterised by bilateral absence or hypoplasia of lower eyelids, macrostomia and multiple other congenital anomalies. Three cases have been reported (McCarthy and West, 1977; Hornblass and Reifler, 1985). In addition to ablepharon and macrostomia, other anomalies common to all patients include auricular deformity, nasal alar deformity, absence of lanugo hair, dry, ichthyotic skin and ambiguous genitalia. A new feature of the syndrome is described--absence of the zygomatic arches. In addition, an expanded and revised classification of the ablepharon macrostomia syndrome and related disorders is presented. Skin graft pigmentation in this black patient has been prevented by prolonged application of sun block.     

A continuous striatal infusion of 6-hydroxydopamine produces a terminal axotomy and delayed behavioral effects

Rat models of Parkinson's disease typically employ a rapid nigral injection of 6-hydroxydopamine (6-OHDA) to produce a near-complete loss of nigrostriatal dopamine neurons, and thus model end stage disease. The present report describes the use of a continuous, low dose infusion of 6-OHDA into the striatum which produces a terminal axotomy of nigrostriatal dopamine neurons and protracted behavioral response. A solution of 6-OHDA in 0.4% ascorbate, delivered at 37°C from osmotic minipumps, was stable for 8 days as determined by its retained toxicity to a dopaminergic neuroblastoma cell line. The continuous infusion of 0.2 μg 6-OHDA per h did not affect the striatal uptake of [³H]GABA, [³H]choline, or [³H]glutamate but reduced [³H]dopamine uptake by 55% within 1.5 days after the start of the infusion. The striatal infusion of 6-OHDA produced a dose-dependent reduction of striatal dopamine and DOPAC levels but did not alter HVA, 5-HT, or 5-HIAA. An increase in amphetamine-induced ipsiversive rotations occurred within 1.5 days after the acute striatal injection of 20 μg or 30 μg of 6-OHDA but required 4 days to develop with the continuous 6-OHDA infusion. The topography of the lesion mapped by [³H]mazindol binding showed that, begining by 1.5 days, a diffuse depletion of terminals encompassed much of the striatum in the 30 μg acute injection group, whereas in the continuously infused rats, the lesion was apparent only by 4 days and was restricted to a smaller and more completely lesioned area. Unlike acutely lesioned animals, continuously infused rats revealed no obvious loss of dopamine neurons in the pars compacta by 5 weeks after 6-OHDA. The continuous striatal infusion of 6-OHDA can produce a topographically limited terminal axotomy of dopamine neurons and a protracted behavioral impairment.     

What has trust got to do with it? Cardiac risk reduction and family physicians' discussions of evidence-based recommendations

The management of patients through the use of evidence-based medicine has become the 'mantra' of medicine within many Western countries. Evidence-based medicine is aimed at providing the best objective, scientific care to all patients, and reducing as far as possible patients' risks of disease and complications from disease. Based on family physicians' discussions of the use of evidence-based recommendations for two cardiac diseases, this paper explores how subjectively-based trust enters into family physicians' decision to use evidence-based medicine. In addition, we show how trust influences physicians' work of recommending evidence-based medicine to patients, and physicians' perceptions of why patients follow recommendations aimed at risk reduction. We conclude that although much of the current discussion about evidence-based medicine assumes a 'rational' model of physician behaviour based on the application of the 'best objective scientific' results, subjectively-based perceptions of trust influence physician practices, and point to the need to understand the power of relational issues in influencing physician practices even when utilizing evidence-based knowledge.