A fluorescence study of the binding of cytochrome C to mixed-phospholipid microvesicles : evidence for a preferred orientation of the bound protein.

Kinetic and equilibrium experiments are reported on the binding of the fluorescent probe 1,8-anilino-naphtalene sulfonate (ANS) to microvesicles of natural lecithin containing 10 per cent of an anionic phospholipip (90 : 10 mixtures). Kinetics discriminated between fast binding to the outer leaflet of the bilayer and apparently slow binding to the inner leaflet controlled by the diffusion of the probe across the bilayer. The equilibrium distribution of ANS between the two leaflets was not dependent on the nature of the anionic species and the spectral properties of bound ANS were identical in all cases investigated. A hyperbolic saturation was observed allowing to propose an affinity scale for the binding of ANS to mixtures of lecithin with phosphatidic acid, phosphatidylinositol, and cardiolipin. The effects on binding of ionic strength and sodium dodecylsulfate were also considered. The binding of horse heart ferricytochrome c to ANS-labelled microvesicles was studied quantitatively making use of the quenching of the probes fluorescence by the heme. Perrin-F?rster energy transfer could be analysed on the basis of a simple model of the physical arrangement of the system which was elaborated from published data referring to ANS and cytochrome c binding to phospholipids. Experimental and theoretical computed values of the quenching efficiency were compared and led to conclude in favor of a preferred orientation of the heme crevice fully accessible from the external space at the lipid interface.     

Block by phencyclidine of acetylcholine and barium induced adrenal catecholamine secretion

Activation of the sympathetic system by phencyclidine (PCP) should result in catecholamine release from the adrenals. However, adrenalectomy does not reduce PCP-induced hypertension. In an attempt to rectify this inconsistency, the direct effects of PCP on the bovine adrenal medulla were examined. At (3×10^?6M), PCP reduced the acetylcholine-(ACh)-induced catecholamine release by 50%. Surprisingly, barium-induced secretion of catecholamines was also reduced by PCP. ACh-induced catecholamine release was not altered by 10^?3M 4-aminopyridine (4 AP), the potassium channel blocker. Thus, calcium antagonist actions of PCP and consequent block of catecholamine secretion from adrenal medulla may explain the lack of effect of adrenalectomy on PCP-induced hypertension. Possible contributions of calcium and/or potassium channel blockade to other manifestations of PCP overdosage are discussed.     

Histochemical study of calcium on T-tubule membranes and in the sarcoplasmic reticulum, in frog twitch muscle fibres at rest and during activity

The trigger calcium hypothesis of signal transmission between T-tubules and terminal cisternae (TC) of the sarcoplasmic reticulum (SR) in twitch muscle fibres implies the presence of calcium along T-tubule membranes at rest and its release upon excitation. To test this hypothesis, calcium was immobilised using a fixing and precipitating solution of glutaraldehyde in phosphate buffer at pH 8.0 and the calcium was substituted for by lead. Simultaneous tension recordings revealed the occurrence of contractions or a burst of twitches upon perfusion with the fixative. Procaine or tetrodotoxin (TTX) was used to inhibit this activity. In fibres without fixative-induced activity, precipitates were observed along T-tubules and in adjoining parts of TC. In activated fibres, tubular and TC precipitates were absent. These results are consistent with the trigger calcium hypothesis. In fibres activated by depolarisation, calcium returned to TC after passing successively through different parts of the SR.     

Reproductive ecology and growth of a population of brown trout (Salmo trutta L.) in an aquifer-fed stream of Old Castile (Spain)

In the River Lobos-Ucero and its tributary the River Avión-Milanos (Duero basin, Old Castile, Central Spain), two limestone streams fed by aquifers, the population of brown trout, as compared with the populations of other European streams, shows a high growth rate, high condition coefficients, short life-span and early age at first maturity. Gonad cycle was also studied. Size distributions of unshed eggs exhibit a dynamic activity with a bi-modal distribution from June onwards, spawning occurred in the last days of November. Fecundity (F) can be predicted from trout length (L, mm) according to the equation: F= –646.47+5.6167 · L. Numbers and standing crop of trout range from 18 to 3903 ind. ha^–1 and 3.6 to 452.9 Kg ha^–1, reaching higher values in the sites close to the aquifers. Egg production had values of 22.4 and 18.0 eggs m^–2 in the Rivers Ucero and Avión-Milanos respectively. Some factors suggested as regulators of these demographical characteristics are discussed in the light of recent literature.