Possible negative ultra-short loop feedback of luteinizing hormone releasing hormone (LHRH) in the ovariectomized rat

To determine if LHRH might act within the brain to modify its own release, repeated blood samples were removed from conscious ovariectomized rats and minute doses of LHRH were injected into the third ventricle (3V). The effect of these injections on plasma LH and FSH was measured by radioimmunoassay (RIA). The higher dose of intraventricular LHRH (10 ng in 2 microliter) induced an increase in plasma LH within 10 min after its injection. Plasma LH decreased for the next 60 min. This was followed by restoration of LH pulses characteristic of the ovariectomized rat. This dose of LHRH slightly elevated plasma FSH concentrations. In stark contrast, a 10 fold lower dose of 1 ng of LHRH injected into the ventricle resulted in a highly significant decrease of plasma LH at 10 min following injection, followed by return of LH pulsations. There was no effect on the pulsatile release of FSH. The results are interpreted to mean that at the higher dose, sufficient LHRH reached the site of origin of the hypophyseal portal vessels in the median eminence so that it diffused into portal vessels and was delivered to the gonadotrophs to induce LH release. In contrast, the lower dose provided sufficient hypothalamic concentrations of the peptide to suppress the discharge of the LHRH neurons, thereby leading to a decline in plasma LH, indicative of an ultrashort-loop negative feedback of LHRH to suppress its own release.     

The effect of the estrous cycle and estrogen on the release of immunoreactive alpha-melanocyte-stimulating hormone

Circulating levels and tissue content of alpha-MSH were measured on the morning of various days of the estrous cycle, and on the afternoon of proestrus in freely moving conscious rats. No surges of alpha-MSH were detected by RIA in the morning of various days of the cycle. The neurointermediate lobe content of alpha-MSH was slightly elevated on diestrus 1 as compared to the levels on diestrus 11 and proestrus but not to estrous levels. No changes in alpha-MSH content were detected in the anterior pituitary, the median eminence, mediobasal hypothalamus and the preoptic area at various stages of the estrous cycle. Plasma alpha-MSH levels were slightly elevated at 1500 hr of proestrus which was followed three hours later by a decline. This profile of plasma alpha-MSH on the afternoon of proestrus was reproduced by the SC administration of estradiol benzoate to long-term ovariectomized rats. These data suggest that, contrary to the results obtained by bioassay of alpha-MSH no surges of alpha-MSH occur on any day of the cycle, although a slight elevation on the afternoon of proestrus was detected. The altered pattern of release of this peptide on the afternoon of proestrus may be induced by estrogen.     

Pancreatic polypeptides affect luteinizing and growth hormone secretion in rats

We have examined the effects of third cerebroventricular (3V) injections of avian and bovine pancreatic polypeptide (APP and BPP) and the C-terminal hexapeptide amide of human PP (CHPP) on the secretion of anterior pituitary hormones in conscious ovariectomized rats. Injection of APP (2.0 micrograms; 472 pmoles) or BPP (5.0 micrograms; 1191 pmoles) decreased plasma levels of luteinizing hormone (LH) when compared to pre-injection levels in these animals or to saline-injected controls. The lower dose of BPP (0.5 micrograms; 119 pmoles) decreased plasma LH versus pre-injection levels and control animals, however, these effects diminished at later times. Plasma growth hormone (GH) also decreased following 3V injections of APP (2.0 micrograms) or BPP (5.0 micrograms). The lower dose of BPP (0.5 microgram) initially inhibited GH release, however, this effect was rapidly reversed and GH levels were significantly greater than those in controls at 60 and 120 min. Injections of BPP or APP did not alter prolactin (PRL) or thyroid stimulating hormone (TSH) secretion. Administration of 2.0 micrograms and 0.2 microgram of CHPP (2488 and 249 pmoles) produced no significant effects on plasma LH, GH, PRL or TSH. APP and BPP had no consistent effects on hormone secretion from dispersed anterior pituitary cells. The results indicate that APP and BPP exert potent central effects which inhibit LH and GH release from the pituitary gland.     

Apparent ornithine decarboxylase activity, measured by 14CO2 trapping, after frozen storage of rat tissue and rat tissue supernatants

Ornithine decarboxylase (ODC) activity of rat tissues was measured by the standard 14CO2 trapping method after frozen storage (-60 or -70 degrees C) of the tissues or their 105,000g supernatants. True ODC activity was determined by two methods: (a) addition of the inhibitors alpha-difluoromethylornithine (DFMO), a specific irreversible inhibitor of ODC, or aminooxyacetate (AOA), an inhibitor that blocks the decarboxylation of ornithine by mitochondrial enzymes; and (b) chromatographic analysis of the reaction products. In the frozen supernatants of liver and spleen, ODC activity changed only slightly after 1 day but increased 29 and 14%, respectively, by 30 days; activity in kidney supernatant decreased 17% after 1 day and remained near that level at 30 days. Kidney and spleen ODC activity was inhibited 90-100% by DFMO, but apparent liver ODC activity was inhibited only 60-75%. In the supernatant prepared from tissue stored frozen for 1 day, apparent ODC activity in liver increased 500% over that activity in the freshly prepared supernatant; at 23 days, apparent activity increased 755% for liver and 121% for kidney. After 23 days, DFMO did not inhibit apparent ODC activity in supernatants from frozen liver and inhibited ODC in frozen kidney by only 49%. With AOA, the ODC activities of the fresh and frozen supernatants were similar, indicating that the large increase in apparent ODC activity in frozen tissue was due to artifacts from the metabolism of ornithine via the mitochondrial pathway. HPLC analysis of the reaction products resulting from the incubation of uniformly labeled [14C]ornithine with the fresh and frozen preparations indicated no increase in putrescine with the frozen preparation.(ABSTRACT TRUNCATED AT 250 WORDS)     

Intergeneric conjugation in Thiobacillus versutus.

In plate matings with Escherichia coli HB101/pUW965::Tn5 (KmR) Thiobacillus versutus reacted as an efficient recipient, producing 10(-2) to 10(-3) kanamycin resistant (KmR) T. versutus exconjugants per donor cell. Analysis of agarose gels of plasmid DNA extracted from the exconjugants confirmed that the suicide vector pUW964 did not persist in the recipient, implying that the kanamycin resistance of the exconjugants is based on effective transposition of Tn5 in T. versutus as well as function of the E. coli kanamycin gene. Transfer was equally efficient when a nalidixate-resistant T. versutus mutant was used as recipient. Hybridization evidence for the presence of Tn5 was consistently negative. The significance of this anomalous result is discussed.     

Ancestral haplotypes reveal the role of the central MHC in the immunogenetics of IDDM

The major histocompatibility complex (MHC) contains multiple and diverse genes which may be relevant to the induction adn regulation of autoimmune responses in insulin dependent diabetes mellitus (IDDM). In addition to HLA class I and II, the possible candidates include TNF, C4, and several other poorly defined polymorphic genes in the central MHC region. This study describes two approaches which take advantage of the fact that the relevant genes are carried by highly conserved ancestral haplotypes such as 8.1 (HLA-B8, TNFS, C4AQO, C4B1, DR3, DQ2). First, three diabetogenic haplotypes (two Caucasoid and one Mongoloid) have been compared and it has been shown that all three share a rare allele of BAT3 as well as sharing DR3, DQ2. In 43 sequential patients with IDDM the cross product ration for BAT3S was 4.8 (p<0.01) and 6.9 for HLA-B8 plus BAT3S (p<0.001). Second, partial or recombinant ancestral haplotypes with either HLA class I (HLA-B8) or II (HLA-DR3, DQ2) alleles were identified. Third, using haplotypic polymorphisms such as the one in BAT3, we have shown that all the patients carrying recombinants of the 8.1 ancestral haplotype share the central region adjacent to HLA-B. These findings suggest that both HLA and non-HLA genes are involved in conferring susceptibility to IDDM, and that the region between HLA-B and BAT3 contains some of the relevant genes. By contrast, similar approaches suggest that protective genes map to the HLA class II region.     

The role of silk in courtship and chemical communication of the false widow spider, <Emphasis Type="Italic">Steatoda grossa</Emphasis> (Araneae: Theridiidae)

In spiders, sex pheromones are often associated with silk produced by females, and function in mate attraction, recognition, and evaluation. Silk-bound pheromones typically elicit courtship behaviour in web-building spiders. Here we (1) describe courtship interactions of Steatoda grossa males with virgin or mated females, and (2) show that silk and methanol extracts of silk produced by virgin females trigger courtship behaviour (silk production) by males, whereas silk of mated females does not. Our results indicate that (1) virgin females produce a silk-bound sex pheromone, (2) males discriminate between virgin and mated females based on silk cues, and (3) male silk likely functions in sexual communication.     

The C-terminal 25 amino acids of the protease and its substrate ICP35 of herpes simplex virus type 1 are involved in the formation of sealed capsids.

The herpes simplex virus type 1 protease and its substrate, ICP35, are involved in the assembly of viral capsids. Both proteins are encoded by a single open reading frame from overlapping mRNAs. The protease is autoproteolytically processed at two sites. The protease cleaves itself at the C-terminal site (maturation site) and also cleaves ICP35 at an identical site, releasing a 25-amino-acid (aa) peptide from each protein. To determine whether these 25 aa play a role in capsid assembly, we constructed a mutant virus expressing only Prb, the protease without the C-terminal 25 aa. Phenotypic analysis of the Prb virus in the presence and absence of ICP35 shows the following: (i) Prb retains the functional activity of the wild-type protease which supports virus growth in the presence of ICP35; (ii) in contrast to the ICP35 null mutant delta ICP35 virus, the Prb virus fails to grow in the absence of ICP35; and (iii) trans-complementation experiments indicated that full-length ICP35 (ICP35 c,d), but not the cleaved form (ICP35 e,f), complements the growth of the Prb virus. The most striking phenotype of the Prb virus is that only unsealed aberrant capsid structures are observed by electron microscopy in mutant-infected Vero cells. Our results demonstrate that the growth of herpes simplex virus type 1 requires the C-terminal 25 aa of either the protease or its substrate, ICP35, and that the C-terminal 25 aa are involved in the formation of sealed capsids.     

Location of cis-acting signals important for RNA encapsidation in the leader sequence of human immunodeficiency virus type 2.

We used a series of deletion mutations in the untranslated leader region of human immunodeficiency virus type 2 (HIV-2) to seek cis-acting packaging signals. Sequences between the 5' major splice donor and the gag initiation codon, where such signals have been identified in HIV-1, appear to make a measurable but very minor contribution to genomic RNA packaging, and deletions here had little effect on viral replication in vitro. Immediately 5' to the splice donor, two regions were identified which, when deleted, caused a significant replication defect. The most proximal of these to the splice donor demonstrated a phenotype consistent with its being a major cis-acting packaging signal in HIV-2.     

Infrared microspectroscopy: Sampling heterogeneity in plant cell wall composition and architecture

The use of probes such as monoclonal and polyclonal antibodies to specific cell wall components, at both the light and electron microscope levels, has demonstrated the diversity in cell wall composition between species, between tissues, between different regions of the cell surface, and even within a single wall. Traditional methods of cell wall analysis have provided valuable information on wall composition and architecture, but, by having to rely on the use of bulk samples, have averaged out this intrinsic heterogeneity. Fourier Transform Infrared (FTIR) microspectroscopy addresses this problem by providing chemical information from an area as small as 10×10 μm of a single cell wall fragment or area of a tissue section that has been imaged with a microscope accessory.
We have used FTIR microspectroscopy as a powerful and extremely rapid assay for wall components and putative cross-links in muro. The spectra are sensitive to polymer conformation, and the use of polarisers in the microscope accessory allows the orientation of particular functional groups to be determined, with respect to the long axis of elongating cells. The spectra constitute species and tissue-specific 'fingerprints', and the use of classical discriminant analysis may provide the opportunity for correlating spectral features with chemical, architectural or rheological wall properties. Spectral mapping of an area of a specimen allows the morphological features resulting from cell growth and differentiation to be characterised chemically at the single cell level.