Pulmonary vasodilatory response to neurohypophyseal peptides in the rat.

Experiments were performed on isolated salt-perfused rat lungs to determine the receptor type(s) responsible for the pulmonary vascular effects of the neurohypophyseal peptides arginine vasopressin (AVP) and oxytocin. Bolus administration of AVP to lungs preconstricted with the thromboxane mimetic U-46619 resulted in a dose-dependent vasodilatory response (approximately 65% reversal of U-46619-induced vasoconstriction at the highest dose tested) that was blocked by pretreatment with a selective V1- but not by a selective V2-vasopressinergic receptor antagonist. Administration of a selective V1-agonist to the preconstricted pulmonary vasculature resulted in a vasodilatory response similar to that observed with AVP (approximately 55% reversal of U-46619 vasoconstriction), which was blocked by prior administration of the selective V1-receptor antagonist. Administration of the selective V2-receptor agonist desmopressin to the preconstricted pulmonary vasculature resulted in a small (approximately 8% reversal of U-46619 vasoconstriction) vasodilatory response that was, nevertheless, greater than that produced by addition of vehicle alone and was attenuated by pretreatment with a selective V2-receptor antagonist. Finally, oxytocin also caused vasodilation in the preconstricted pulmonary vasculature; however, the potency of oxytocin was approximately 1% of AVP, and the vasodilation produced by oxytocin was blocked by prior administration of a selective V1-receptor antagonist, suggesting that oxytocin acts via V1-vasopressinergic receptor stimulation. We conclude from these experiments that AVP and oxytocin dilate the preconstricted pulmonary vasculature primarily via stimulation of V1-vasopressinergic receptors. V2-receptor stimulation results in a minor vasodilatory response, although its physiological significance is unclear.     

Phosphorylation site of eukaryotic initiation factor 4E

Eukaryotic protein synthesis initiation factor 4E (eIF-4E) was labeled in situ with [32P]orthophosphate in cultured HeLa cells and rabbit reticulocytes and purified by affinity chromatography. Tryptic digestion yielded one labeled peptide which contained predominantly serine and lysine. After treatment of the protein with citraconic anhydride to block epsilon-amino groups of lysyl residues, tryptic digestion yielded a labeled peptide whose composition was consistent with the structure Trp-Ala-Leu-Trp-Phe-Phe-Lys-Asn-Asp-Lys-Ser(P)-Lys-Thr-Trp-Gln-Ala-Asn-L eu-Arg, one of the arginyl peptides predicted from the human eIF-4E cDNA sequence. The only serine in this peptide is located at position 53 of eIF-4E. Thus, it is concluded that eIF-4E contains a single site of phosphorylation for an endogenous protein kinase, which is Ser-53 in the human eIF-4E sequence.     

Ethanol exhibits alpha receptor blocking-like properties in anesthetized rats

The ability of ethanol to reduce alpha-adrenergic receptor-mediated pressor responsiveness in vivo was investigated in chloralose-anesthetized male Sprague-Dawley rats. Catheters were inserted in the jugular vein and the femoral artery of rats for the injection of drugs and the measurement of blood pressure, respectively. Dose-response curves for phenylephrine and norepinephrine were constructed by plotting the change in mean arterial pressure following a bolus dose of the agent against the dose of the pressor agent used. Following construction of an initial dose-response curve, animals were challenged with either a 1 g/kg dose of ethanol or an equivalent volume of saline (iv) and the dose-response curves were repeated. Using a similar protocol, pressor responsiveness was evaluated in animals pretreated with either yohimbine (1 mg/kg) or prazosin (3.9 micrograms/kg), a dose sufficient to produce partial blockade of alpha receptor-mediated pressor responsiveness, and then treated with ethanol. Ethanol produced a partial blockade of alpha receptors when the animals were challenged with either phenylephrine or norepinephrine. This blockade produced by ethanol was shown to be similar to that produced by the receptor blocking agents used in this study. To rule out any nonspecific effects of ethanol in reducing vascular reactivity, some animals were challenged with angiotensin II both before and after treatment with ethanol, yohimbine, or prazosin and after both drugs were administered together. Ethanol, as well as the alpha 1- and alpha 2-adrenergic blocking agents tested failed to have any significant effect on angiotensin II-pressor responsiveness, ruling out any nonspecific effect of ethanol on the vasculature. It is concluded, therefore, that ethanol has alpha receptor blocking-like activity in vivo.     

Brefeldin A redistributes resident and itinerant Golgi proteins to the endoplasmic reticulum

Brefeldin A (BFA) has been reported to block protein transport from the ER and cause disassembly of the Golgi complex. We have examined the effects of BFA on the transport and processing of the vesicular stomatitis virus G protein, a model integral membrane protein. Delivery of G protein to the cell surface was reversibly blocked by 6 micrograms/ml BFA. Pulse-label experiments revealed that in the presence of BFA, G protein became completely resistant to endoglycosidase H digestion. Addition of sialic acid, a trans-Golgi event, was not observed. Despite processing by cis- and medial Golgi enzymes, G protein was localized by indirect immunofluorescence to a reticular distribution characteristic of the ER. By preventing transport of G protein from the ER with the metabolic inhibitor carbonyl cyanide m-chlorophenylhydrazone or by use of the temperature-sensitive mutant ts045, which is restricted to the ER at 40 degrees C, we showed that processing of G protein occurred in the ER and was not due to retention of newly synthesized Golgi enzymes. Rather, redistribution of preexisting cis and medial Golgi enzymes to the ER occurred as soon as 2.5 min after addition of BFA, and was complete by 10-15 min. Delivery of Golgi enzymes to the ER was energy dependent and occurred only at temperatures greater than or equal to 20 degrees C. BFA also induced retrograde transport of G protein from the medial Golgi to the ER. Golgi enzymes were completely recovered from the ER 10 min after removal of BFA. These findings demonstrate that BFA induces retrograde transport of both resident and itinerant Golgi proteins to the ER in a fully reversible manner.     

Order of bovine DRB3, DYA,and PRL determined by sperm typing

The order and recombination fractions () between the bovine major histocompatibility complex DRB3, DYA, and prolactin (PRL) genes were determined by typing of 254 sperm from a triply heterozygous bull. A recently developed method, primer extension preamplification (PEP), was used to amplify the bovine sperm genome prior to amplification of specific loci by the polymerase chain reaction (PCR). At least 28 copies of the DRB3, PRL, or DYA gene were obtained from 50 cycles of PEP. For sperm typing, alleles of each locus were discriminated by restriction endonuclease cleavage of PCR products and polyacrylamide gel electrophoresis of the restriction fragments. The most likely gene order is PRL-DRB3-DYA, with =0.025 (±0.012) and =0.150 (±0.024), respectively. The odds are 128:1 in favor of this order in comparison with the second most likely order DRB3-PRL-DYA. Our results demonstrate the power of sperm typing in concert with PEP for multilocus gene mapping.     

Influenza virus hemagglutinin trimers and monomers maintain distinct biochemical modifications and intracellular distribution in brefeldin A-treated cells.

Brefeldin A (BFA) induces the retrograde transport of proteins from the Golgi complex (GC) to the endoplasmic reticulum (ER). It is uncertain, however, whether the drug completely merges the ER with post-ER compartments, or whether some of their elements remain physically and functionally distinct. We investigated this question by the use of monoclonal antibodies specific for monomers and trimers of the influenza virus hemagglutinin (HA). In untreated influenza virus-infected cells, monomers and trimers almost exclusively partition into the ER and GC, respectively. In BFA-treated cells, both monomers and trimers are detected in the ER by immunofluorescence. Cell fractionation experiments indicate, however, that whereas HA monomers synthesized in the presence of BFA reside predominantly in vesicles with a characteristic density of the ER, HA trimers are primarily located in lighter vesicles characteristic of post-ER compartments. Biochemical experiments confirm that in BFA-treated cells, trimers are more extensively modified than monomers by GC-associated enzymes. Additional immunofluorescence experiments reveal that in BFA-treated cells, HA monomers can exist in an ER subcompartment less accessible to trimers and, conversely, that trimers are present in a vesicular compartment less accessible to monomers. These findings favor the existence of a post-ER compartment for which communication with the ER is maintained in the presence of BFA and suggest that trimers cycle between this compartment and the ER, but have access to only a portion of the ER.     

Overgrowth in a marine epifaumal community: Competitive hierarchies and competitive networks

Summary The frequencies with which organisms of a species overgrew or were overgrown by organisms of other species in a marine epifaunal community were estimated. The ranking of the ability of the major taxonomic groups to overgrow others was basically hierarchical:ascidianssponges>bryozoans>barnacles, polychaetes, tubicolous amphipods, hydroids. In contrast, the ranking of the competitive ability of species in the community did not form a simple linear hierarchy and there was no single competitively dominant species (measured in terms of overgrowth). There were often no significant differences in the ability of species to overgrow each other within the three major taxonomic groups of sponges, ascidians and bryozoans. Such results were common also between the species of large sponges and ascidiams which dominated substrata immersed for periods longer than two years.A lack of a significant difference in the competitive ability of species was usually the result of (a) frequent formation of delay/ties or standoffs and (b) changes in the outcome of interactions due to change in the relative size of interacting colonies. In many two-species interactions the species which had the larger colony in a given encounter had a greater probability of winning.When the range of colony sizes of two species was similar there was often no significant difference between the competitive ability of each species. Such cases without a clearcut winner often represented a backloop in an otherwise hierarchical sequence of competitive ability, i.e. Species A beats Species B, Species B beats Species C, no significant differences in competitive ability between Species C and A. No examples of competitive networks of the form Species A beats Species B, Species B beats Species C, Species C beats Species A were found. Backloops in otherwise hierarchical sequences (no significant differences in competitive ability) occurred most frequently between species within the same major taxonomic groups and were the result of a very even balance in the generalised competitive mechanism of overgrowth.It seems probable that backloops in hierarchical sequences are more commonly due to the absence of clear competitive dominance in interactions between species (reversals in the outcome of overgrowth interactions and standoffs), rather than to direct backloops formed by a specialised or to a generalised competitive mechanism. Network-like arrangements of competitive ability formed by the type of processes described here are likely to contribute significantly to the high levels of species diversity observed in many marine epifaunal communities subject to low levels of physical disturbance.