Scrotal heating stimulates panting and reduces body temperature similarly in febrile and non-febrile rams (Ovis aries)

It is known that heating the ram scrotum stimulates heat loss resulting in a decrease in body temperature and that during fever core temperature increases, but local scrotal thermoeffectors operate to maintain normal scrotal temperature. We have investigated whether scrotal warming influences core body temperature and the panting effector during fever generation. We measured rectal temperature, intrascrotal temperature, scrotal skin temperature and respiratory frequency in four adult Merino rams following intravascular injection of saline or lipopolysaccharide at an ambient temperature of 18-20 degrees C while scrotal skin temperature was maintained at 33 degrees C or elevated to 41 degrees C. Compared to maintaining normal scrotal temperature, heating the scrotum increased respiratory frequency and reduced rectal temperature by a similar amount following LPS as following saline. Fever was associated with decreased respiratory frequency compared to saline at both 33 and 41 degrees C scrotal temperature, suggesting that the fever was generated mainly by decreasing respiratory heat loss. We conclude that scrotal thermal afferent stimulation resulted in an offset for the set-point of body temperature regulation in both normothermic and febrile rams.     

Influence of oral contraceptive use on endothelial t-PA release in healthy premenopausal women

We determined the influence of oral contraceptives (OC) on the capacity of the endothelium to release tissue-type plasminogen activator (t-PA). Twenty-three healthy premenopausal women were studied: 12 nonusers and 11 users of OC. Net endothelial release rates of t-PA were calculated as the product of the arteriovenous concentration gradient and forearm plasma flow in response to intra-arterial bradykinin (BK: 12.5-50 ng. 100 ml tissue(-1) x min(-1)) and sodium nitroprusside (SNP: 1.0-4.0 microg x 100 ml tissue(-1) x min(-1)). Net release of t-PA antigen and increment in t-PA activity across the forearm to BK increased (P < 0.01) in a dose-dependent fashion and to similar extents in the nonusers and users of OC. At the highest BK dose, net release of t-PA antigen was 64.5 +/- 8.2 and 66.2 +/- 15.4 ng x 100 ml tissue(-1) x min(-1) in the nonusers and users of OC, whereas the net increment in t-PA activity was 18.6 +/- 3.0 and 16.0 +/- 2.0 IU. 100 ml tissue(-1) x min(-1), respectively. There was no effect of SNP on t-PA release in either group. These results indicate that endothelial t-PA release is not altered in premenopausal women who use oral contraception.     

Acute phase cytotoxic T lymphocyte escape is a hallmark of simian immunodeficiency virus infection

Cytotoxic T-lymphocyte (CTL) responses peak coincident with the decline in acute HIV viremia. Despite two reports of CTL-resistant HIV variants emerging during acute infection, the contribution of acute CTL escape to HIV pathogenesis remains unclear. Difficulties inherent in studying acute HIV infection can be overcome by modeling virus-host interactions in SIV-infected rhesus macaques. We sequenced 21 complete simian immunodeficiency virus (SIV)mac239 genomes at four weeks post-infection to determine the extent of acute CTL escape. Here we show that viruses from 19 of 21 macaques escaped from CTLs during acute infection and that these escape-selecting CTLs were responsive to lower concentrations of peptide than other SIV-specific CTLs. Interestingly, CTLs that require low peptide concentrations for stimulation (high 'functional avidity') are particularly effective at controlling other viral infections. Our results suggest that acute viral escape from CTLs is a hallmark of SIV infection and that CTLs with high functional avidity can rapidly select for escape variants.     

Proteomic analysis of the proliferative and secretory phases of the human endometrium: protein identification and differential protein expression

Proteomic analyses of the proliferative and secretory phases of the human endometrium were carried out to identify proteins and discover differentially expressed proteins using isotope-coded affinity tags, three stages of chromatographic separation and online tandem mass spectrometry (MS/MS). From an initial list of 346 proteins identified by ProICAT, manual inspection of MS/MS spectra and confirmatory searches pared the list down to 119 positively identified proteins. Only five of the proteins showed consistent differential expression. The utility of some of these proteins as indicators of true differential expression in the endometrium is open to discussion. The two proteins with unquestionable differential expressions in the secretory endometrium are: glutamate NMDA receptor subunit zeta 1 precursor and FRAT1. Some of the proteins that show no differential expression have previously been examined in gene-expression studies with similar conclusions.     

Purification and some properties of human placental glucose-6-phosphate dehydrogenase

Glucose-6-phosphate dehydrogenase was purified from human placenta using DEAE-Sepharose fast flow, 2',5'-ADP Sepharose 4B column chromatography, and chromatofocusing on PBE 94 with PB 74. The enzyme was purified with 62% yield and had a specific activity of 87 units per milligram protein. The pH optimum was determined to be 7.8, using zero buffer extrapolation method. The purified placental glucose-6-phosphate dehydrogenase gave two activity bands on native PAGE: one band, constituting about 3--5% of total activity, moved slower than the remaining 95%. Among the activity bands only the faster moving band gave a band on protein staining. The slower moving band, which probably corresponded to the higher polymeric form of the G6PD with high specific activity, was not seen on native PAGE due to insufficient protein for Coomassie brilliant blue staining. The observation of one band on SDS--PAGE with an M(r) of 54 kDa and a specific activity lower than expected, suggests the presence of both forms of the G6PD, the high polymeric form at low concentration and the inactive form at high concentration, in our preparation. Measuring the activities of placental glucose-6-phosphate dehydrogenase between 20 and 50 degrees C, the activation energy, activation enthalpy, and Q(10) were calculated to be 8.16 kcal/mol, 7.55 kcal/mol, and 1.57, respectively. It was found that human placental G6PD obeys Michaelis-Menten kinetics. K(m) values were determined using the concentration ranges of 20--300 microM for G6P and 10--200 microM for NADP(+). The K(m) value for G6P was 40 microM; the K(m) value NADP(+) was found to be 20 microM. Double-reciprocal plots of 1/Vm vs 1/G6P (at constant [NADP(+)]) and of 1/Vm vs 1/NADP(+) (at constant [G6P]) intersected at the same point on the 1/V(m) axis to give V(m) = 87 U/mg protein.     

New sample preparation method for the capillary electrophoretic determination of adenylate energy charge in human erythrocytes

The first step in the separation of adenine nucleotides from different types of tissues or cells is deproteinization. Several sample preparation methods successfully used for a number of tissues or cells failed to work with erythrocytes. Use of strong acids or bases for deproteinization resulted in a low yield due to the hydrolysis of adenine nucleotides. Moreover, the neutralization of these acids or bases increased the ionic strength, resulting in broad and overlapping peaks. In neutral salt precipitation methods, saturated salts caused clogging of the capillaries. A new deproteinization procedure method was developed. The samples were deproteinized by heating of erythrocytes in boiling distilled water at 95 degrees C for 5 min. The denatured proteins were removed by centrifugation and membrane filtration. The adenine nucleotides were then separated using a polyacrylamide coated capillary. Depending on the type, diameter, length of the capillary and the voltage applied, an average of 16.50 min was sufficient for the separation of adenine nucleotides. All adenine nucleotides were clearly resolved and gave very sharp peaks. The amount of each adenine nucleotide was calculated from the areas under the peaks and AEC values were calculated using the integrator software. The AEC value of 0.91+/-0.04 (n=10) obtained for healthy persons was in good agreement with the literature value of 0.85-0.95. These reported method for sample preparation and capillary electrophoresis is simple, fast and inexpensive compared to the previously reported sample preparation, HPLC and enzymatic methods for the determination of AEC.     

Correlating Behavioral Responses to fMRI Signals from Human Prefrontal Cortex: Examining Cognitive Processes Using Task Analysis

The aim of this methods paper is to describe how to implement a neuroimaging technique to examine complementary brain processes engaged by two similar tasks. Participants'' behavior during task performance in an fMRI scanner can then be correlated to the brain activity using the blood-oxygen-level-dependent signal. We measure behavior to be able to sort correct trials, where the subject performed the task correctly and then be able to examine the brain signals related to correct performance. Conversely, if subjects do not perform the task correctly, and these trials are included in the same analysis with the correct trials we would introduce trials that were not only for correct performance. Thus, in many cases these errors can be used themselves to then correlate brain activity to them. We describe two complementary tasks that are used in our lab to examine the brain during suppression of an automatic responses: the stroop¹ and anti-saccade tasks. The emotional stroop paradigm instructs participants to either report the superimposed emotional ''word'' across the affective faces or the facial ''expressions'' of the face stimuli^1,2. When the word and the facial expression refer to different emotions, a conflict between what must be said and what is automatically read occurs. The participant has to resolve the conflict between two simultaneously competing processes of word reading and facial expression. Our urge to read out a word leads to strong ''stimulus-response (SR)'' associations; hence inhibiting these strong SR''s is difficult and participants are prone to making errors. Overcoming this conflict and directing attention away from the face or the word requires the subject to inhibit bottom up processes which typically directs attention to the more salient stimulus. Similarly, in the anti-saccade task^3,4,5,6, where an instruction cue is used to direct only attention to a peripheral stimulus location but then the eye movement is made to the mirror opposite position. Yet again we measure behavior by recording the eye movements of participants which allows for the sorting of the behavioral responses into correct and error trials⁷ which then can be correlated to brain activity. Neuroimaging now allows researchers to measure different behaviors of correct and error trials that are indicative of different cognitive processes and pinpoint the different neural networks involved.     

Mutual Use of Trail-Following Chemical Cues by a Termite Host and Its Inquiline

Termite nests are often secondarily inhabited by other termite species ( = inquilines) that cohabit with the host. To understand this association, we studied the trail-following behaviour in two Neotropical species, Constrictotermes cyphergaster (Termitidae: Nasutitermitinae) and its obligatory inquiline, Inquilinitermes microcerus (Termitidae: Termitinae). Using behavioural experiments and chemical analyses, we determined that the trail-following pheromone of C. cyphergaster is made of neocembrene and (3Z,6Z,8E)-dodeca-3,6,8-trien-1-ol. Although no specific compound was identified in I. microcerus, workers were able to follow the above compounds in behavioural bioassays. Interestingly, in choice tests, C. cyphergaster prefers conspecific over heterospecific trails while I. microcerus shows the converse behaviour. In no-choice tests with whole body extracts, C. cyphergaster showed no preference for, while I. microcerus clearly avoided heterospecific trails. This seems to agree with the hypothesis that trail-following pheromones may shape the cohabitation of C. cyphergaster and I. microcerus and reinforce the idea that their cohabitation is based on conflict-avoiding strategies.     

Multiple splicing variants of cdc25B regulate G2/M progression.

Progression through G2 phase into mitosis is regulated by the activation of the mitotic cyclin/cdk complexes, which are in turn activated cdc25B and cdc25C phosphatases. Here we report that alternate splicing produces at least five variants of cdc25B, although only cdc25B2 and cdc25B3 are detectable as proteins. Analysis of these two variants shows that cdc25B2 is expressed at lower levels relative to cdc25B3 in all cell lines tested, and the expression of both increased markedly during G2 and mitosis. Overexpression of the catalytically inactive version of either cdc25B variant produced a G2 arrest implicating both in regulating G2/M progression.