Characterization of high affinity binding sites for charybdotoxin in sarcolemmal membranes from bovine aortic smooth muscle. Evidence for a direct association with the high conductance calcium-activated potassium channel

Charybdotoxin (ChTX), a peptidyl inhibitor of the high conductance Ca2+-activated K+ channel (PK,Ca), has been radiolabeled to high specific activity with 125I, and resulting derivatives have been well separated. The monoiodotyrosine adduct blocks PK,Ca in vascular smooth muscle with slightly reduced potency compared with the native peptide under defined experimental conditions. [125I]ChTX, representing this derivative, binds specifically and reversibly to a single class of sites in sarcolemmal membrane vesicles prepared from bovine aortic smooth muscle. These sites display a Kd of 100 pM for the iodinated toxin, as determined by either equilibrium or kinetic binding analyses. Binding site density is about 500 fmol/mg of protein in isolated membranes. The addition of low digitonin concentrations to disrupt the vesicle permeability barrier increases the maximum receptor concentration to 1.5 pmol/mg of protein, correlating with the observations that ChTX binds only at the external pore of PK,Ca and that the membrane preparation is of mixed polarity. Competition studies with ChTX yield a Ki of about 20 pM for native toxin. Binding of [125I]ChTX is modulated by ionic strength as well as by metal ions that are known to interact with PK,Ca. Moreover, tetraethylammonium ion, which blocks PK,Ca with moderately high affinity when applied at the external membrane surface, inhibits [125I]ChTX binding in an apparently competitive fashion with a Ki similar to that found for channel inhibition. In marked contrast, agents that do not inhibit PK,Ca in smooth muscle (e.g. tetrabutylammonium ion, other toxins homologous with ChTX, and pharmacological agents that modulate the activity of dissimilar ion channels) have no effect on [125I]ChTX binding in this tissue. Taken together, these results suggest that the binding sites for ChTX which are present in vascular smooth muscle are directly associated with PK,Ca, thus identifying [125I]ChTX as a useful probe for elucidating the biochemical properties of these channels.     

Exploring the Potential of Laser Ablation Carbon Isotope Analysis for Examining Ecology during the Ontogeny of Middle Pleistocene Hominins from Sima de los Huesos (Northern Spain)

Laser ablation of tooth enamel was used to analyze stable carbon isotope compositions of teeth of hominins, red deer, and bears from middle Pleistocene sites in the Sierra de Atapuerca in northern Spain, to investigate the possibility that this technique could be used as an additional tool to identify periods of physiological change that are not detectable as changes in tooth morphology. Most of the specimens were found to have minimal intra-tooth variation in carbon isotopes (< 2.3‰), suggesting isotopically uniform diets through time and revealing no obvious periods of physiological change. However, one of the two sampled hominin teeth displayed a temporal carbon isotope shift (3.2‰) that was significantly greater than observed for co-occurring specimens. The δ¹³C value of this individual averaged about -16‰ early in life, and -13‰ later in life. This isotopic change occurred on the canine crown about 4.2 mm from the root, which corresponds to an approximate age of two to four years old in modern humans. Our dataset is perforce small owing to the precious nature of hominid teeth, but it demonstrates the potential utility of the intra-tooth isotope profile method for extracting ontogenetic histories of human ancestors.     

Luteal phase defects: a possible relationship between short hyperthermic phase and sporadic sexual behavior in women

Sporadic (less than regular weekly) coital activity is associated with a particular type of infertility—the short hyperthermic (luteal) phase. This preliminary report is the first suggestion that a reduced frequency of human sexual activity might, itself, be associated with subfertile reproductive function.     

The susceptibility of the moderate halophile Vibrio costicola to heavy metals

Fifty-eight strains of the moderate halophile Vibrio costicola , including both culture collection strains and freshly isolated strains from solar salterns, were examined for their susceptibility to 10 heavy metal ions by using an agar dilution technique. All strains were sensitive to cadmium, copper, silver, zinc and mercury. This latter ion showed the highest activity even at 0·05 mmol/1 metal concentration. On the other hand, all strains were similarly tolerant to lead, and a great proportion of them were also tolerant to nickel (91%) and chromium (88%). Only 44% and 14% of them showed tolerance to arsenate and cobalt, respectively. The majority of strains (96·4%) were multiply metal-tolerant, with three different metal ion tolerances as the major pattern.     

Phenol oxidases from Rhodnius prolixus: Temporal and tissue expression pattern and regulation by ecdysone

Rhodnius prolixus 5th instar nymphs have significant PO enzymatic activity in the anterior midgut, fat body and hemolymph. The tissue with the major amount of PO activity is the anterior midgut while those with higher specific activities are the fat body and hemolymph. In this work the temporal pattern of PO enzymatic activity in different tissues was investigated. In fat body, PO peaks occur at 7, 12 and 16 days after a blood meal. In hemolymph, PO diminishes until day 7, and then recovers by day 14. In the anterior midgut tissue, PO peaks on day 9 and just before ecdysis; a similar pattern was observed in the anterior midgut contents. Some of these activities are dependent on the release of ecdysone, as feeding blood meal containing azadirachtin suppresses them and ecdysone treatment counteracts this effect. These results suggest that during the development of the 5th instar, the insect has natural regulating cycles of basal PO expression and activation, which could be related to the occurrence of natural infections. The differences in temporal patterns of activity and the effects of azadirachtin and ecdysone in each organ suggest that, at least in R. prolixus, different tissues are expressing different PO genes.     

Functional architecture of the ligamentum arteriosum in adults]

The ligamentum arteriosum was studied in eight normal adult human hearts obtained at autopsy. Four of these hearts were treated by the Zemper method and extensively dissected under a steromicroscope (from X 6 up to X 40); two were embedded in celloidin and sectioned at 100mugm; two were embedded in paraffin and sectioned one 50mugm and the other at 75 mugm. Section were stained by the axan and the resorcin-fuchsin methods. Unstained sections were examined under polarized light.The ligamentum arteriosum may be considered as a small smooth muscle, whose origin is in the arteria pulmonalis sinistra and the insertion in the arcus aortae. Therefore, the ligamentum arterisum may be considered as a myoelastic system, included in a collagenous stroma. Muscle, elastic and collagen fibers present a cross-spiral disposition. Being a myoelastic system with a collagen component, the ligamentum arteriosum cannot be considered as a simple arcus aortae support; it must play some functional role in controlling the arcus aortae curvature during the several different steps of the cardiac cycle.     

Ultrastructural changes in the renal filtration barrier during hibernation of the common dormouse (Eliomys quercinus L.)

We have studied kidney samples of 16 garden dormouses (Eliomys quercinus L.) divided into two groups, 8 hibernating and 8 non-hibernating. Hibernation produces structural modifications in the glomerular ultrafilter: (1) the endothelial pores decrease in number and size; (2) the podocytic food processes increase in number and the slit pores decrease in size; (3) in the basement membrane there are no structural morphological modifications. In short, we could say that the permeability of the glomerular ultrafilter decreases during hibernation. This fact helps to understand the decrease in the rate of urine formation that takes place in the presence of a low body temperature and a metabolic depression.