The size of the extracellular space in the isolated midgut ofManduca sexta

The size of the extracellular space in the isolated midgut ofManduca sexta measured statically with labeled sucrose and with labeled inulin was 48 and 50 percent of tissue water respectively. Both labeled sulfate and labeled mannitol yielded loading spaces which approached the volume of the tissue water with increasing pulse time and are not valid markers of the extracellular space. A washout method yielded unreliable values for the sucrose and inulin spaces and its use cannot be justified using presently available midgut chamber designs. Values of the sucrose space measured statistically in this study and other studies using different Lepidopteran larvae and different perfusion chambers range from 42 to 48 percent of the tissue water. Agreement between published values of the sulfate space and these values of the sucrose space is fortuitous, the labeling pulses being short enough to yield approximately half saturation of the actual sulfate loading space.     

Early events in neuromuscular junction formation in vitro: induction of acetylcholine receptor clusters in the postsynaptic membrane and morphology of newly formed synapses

The development of clusters of acetylcholine (ACh) receptors at newly formed synapses between embryonic chick spinal cord and muscle cells grown in vitro has been studied by iontophoretic mapping with ACh. A semi-automated technique using on-line computer analysis of ACh responses and a photographic system to record the position of each ACh application permit the rapid construction of extensive and detailed maps of ACh sensitivity. Clusters of receptors, evident as peaks of ACh sensitivity, are present on many uninnervated myotubes. The distribution of ACh sensitivity closely parallels the distribution of 125I-alpha-bungarotoxin binding sites on the same muscle cell. In all cases where individual myotubes were adequately mapped before and after synapse formation, ingrowing axons induced new clusters of receptors rather than seeking out preexisting clusters. Synapses can form at active growth cones within 3 h of nerve-muscle contact. New receptor clusters can appear beneath neurites within a few hours. Many of the uninnervated clusters on innervated myotubes disappear with time. In contrast, receptor clusters on uninnervated myotubes remain in the same location for many hours. Synaptic clusters and clusters on uninervated myotubes are stable even though individual receptors are metabolized rapidly. The morphology of several identified sites of transmitter release was examined. At the scanning EM level, synapses appeared as small, rough-surfaced varicosities with filopodia that radiated outwards over the muscle surface. One synapse was studied by transmission EM. Acetylcholinesterase and a basement lamina were present within the synaptic cleft.     

Characteristics of anion transport in cat and dog red blood cells

Summary Self-exchange of chloride and sulfate in dog and cat red cells has been measured under equilibrium conditions. The rates of efflux for these anions are approximately twofold higher in dog compared to cat red blood cells. Although the rates differ, the anion exchange systems of these two red cell types exhibit many common properties. The dependence of³⁵SO₄ efflux on the intracellular SO₄ concentration, the pH dependence and the inhibition of³⁵SO₄ efflux by Cl and SITS are almost identical in dog and cat red cells. Nystatin treatment was used to study the dependence of³⁶Cl efflux on internal Cl. Chloride efflux exhibits saturation in both cell types with dog red cells possessing a higherV _max andK _1/2 than cat red cells. The number of anion transport sites was estimated by extrapolation to the number of molecules of dihydro DIDS (H₂DIDS, where DIDS is 4,4-diisothiocyano-2,2 stilbene-disulfonic acid) which were bound at 100% inhibition of transport. The results indicate that either the turnover numbers for anion transport differ in dog, cat, and human red cells or that there is heterogeneity in the function of the membrane components which bind H₂DIDS.     

Mutations affecting the formation of acetohydroxy acid synthase II in Escherichia coli K-12.

Summary Genetic mapping experiments have established that two recently isolated valine-resistant mutants of the K-12 strain of Escherichia coli have lesions lying between ilvE and rbs. These lesions allowed expression of the ilvG gene, specifying the valine-insensitive acetohydroxy acid synthase (synthase II) and an increased expression of the ilvEDA operon. In this respect, they resembled an earlier described ilvO lesion that was reported to lie between ilvA and ilvC. All three lesions were cis-dominant in cis-trans tests. Reexamination of the earlier studied ilvO lesion revealed that it, too, lies between ilvE and rbs. Valine-sensitive derivatives with lesions presumed to be in ilvG were selected from each of the valine-resistant strains. In two of the valine-resistant strains, the ilvG mutations were on the rbs side of ilvO, indicating a gene order rbs-ilvG-ilvO-ilvE-ilvD-ilvA-ilvC. In one of the recently isolated valine-resistant stocks, however, the apparent ilvG mutation was found to be between ilvE and the aline resistance marker. This finding suggests that either ilvO and ilvG mutations are interspersed or there is another locus, ilvR, that behaves phenotypically like ilvO and which lies between ilvG and rbs.     

CORRECTION OF THE NAME PSEUDOTETRAÉDRON POLYMORPHUM (CHLOROPHYCEAE)1

A unicellular chlorococcalean alga, Pseudotetraëdron polymorphum MacEntee, Bold & Archibald is renamed Chlorotetratëdron polymorphum (MacEntee, Bold & Archibald) MacEntee, Bold & Archibald became the former generic designation has been preempted by a xanthophycean alga.     

An inhibitor of dopamine uptake,LR5182, CIS-3-(3,4-dichlorophenyl)-2-N,N-dimethylaminomethyl-bicyclo-[2,2,2]-octane,hydrochloride

LR5182 inhibited the uptake of dopamine in rat striatal synaptosomes and the uptake of norepinephrine in cortical synaptosomes with inhibitor constants, Ki values, of 3nM and 58nM, respectively. It was only a week inhibitor of serotonin uptake in cortical synaptosomes with a Ki value of 1.7μM. The uptake of dopamine and norepinephrine were significantly lowered within an hour after an intraperitoneal injection of LR5182. Among known inhibitors of dopamine uptake in synaptosomes of rat brain, LR5182 is most effective and selective. The rigid structure of LR5182 (Figure 1) suggested a gauche conformation of dopamine to be favored by the striatal uptake of dopamine.     

A study of prostaglandin F2α as the luteolysin in swine: III effects of estradiol valerate on prostaglandin F, progestins, estrone and estradiol concentrations in the utero-ovarian vein of nonpregnant gilts

Polyvinyl catheters were placed into the right and left utero-ovarian veins and saphenous vein and artery of three control (C) and four estradiol valerate (EV) treated gilts on Day 9 after onset of estrus. The EV treated gilts received 5mg EV/day on Days 11 through 15 after onset of estrus. On Days 12 through 17 utero-ovarian vein blood samples were collected at 15 min intervals from 0700 to 1000 hr and 1900 to 2200 hr and single samples were taken at 1100 and 2300 hr. Peripheral blood samples (saphenous vein or artery) were taken at 0700, 1100, 1900 and 2300 hr from Day 12 until the control gilts returned to estrus or until Day 25 for EV treated gilts and used to measure plasma steroid hormone concentrations. Utero-ovarian vein prostaglandin F (PGF) concentrations (ng/ml, n=1,177) were measured by RIA. Status (control EV treated gilts) by day interactions were detected (P=.10). Curvilinear day trends were detected for plasma PGF concentrations in control (P<.01) but not EV treated gilts. PGF concentrations ( ) for control and EV treated gilts were 1.20 ± 2.08 and .26 ± .84 ng/ml, respectively. PGF peaks (concentrations greater than + 2 S.D.) occured with greater frequency in control gilts (X² = 4.87; P<.05). The interestrus interval ( ) for control and treated gilts was 19.0 ± .6 and 146.5 ± 74.8 days, respectively. Data indicate that estradiol valerate may exert its luteotrophic effect by preventing PGF release from the uterus.     

Continuous titration of difference absorption upon binding of 4-methylumbelliferyl glycosides to concanavalin A and peanut agglutinin

A continuous titration of absorption differences is described. Equal volumes of the titration fluid are dispensed from two micrometer-driven Hamilton gas-tight syringes into two 1 × 1 × 4.5-cm cuvettes. These are placed in the reference and sample beam. Each cuvette stopper is equipped with a capillary inlet connected to a syringe and with a minimotor for continuous stirring. Details of the stirring device are given. The delivered volumes of titration fluid are sufficiently reproducible to allow titration of absorption differences as a function of chromophore concentration. The usefulness of this approach is tested with the binding of 4-methylumbelliferyl α-d-mannopyranoside and concanavalin A as a well-characterized system. It is applied to the binding of similarly labeled anti-t disaccharide with the lectin from peanuts. With both lectins, the change in molecular extinction coefficient of the ligand and the association constant, valid for the entire protein saturation range, were obtained. The results are identical to those from other methods, including equilibrium dialysis.     

Evidence for two conformational forms of monhistone protein BA which differ in their affinity for DNA

Nonhistone protein BA_free was purified from the 0.075 M NaCl/0.025 M $\text{EDTA}\text{pH}$ 8 extract of whole rat liver nuclei while protein BA_bound was isolated from the 0.05 M Na₂HPO₄/8 M urea/1% β-mercaptoethanol/pH 7.6 extract of dehistonized rat liver chromatin. Chromatin associated protein BA_bound was able to bind 60% of the [³H] DNA in a nitrocellulose filter binding assay while nucleoplasmic protein BA_free showed essentially no DNA binding activity. Circular dichroism analysis of the two forms of protein BA revealed substantial differences in their conformations. Protein BA_free was found to have an α-helix content of 41% while protein BA_bound displayed a spectrum more typical of unordered or β-turn structures.