An experimental test of a model for repeated Ca2+ spikes in osteoblastic cells.

A model for cytosolic Ca2+ spikes is presented that incorporates continual influx of Ca2+, uptake into an intracellular compartment, and Ca(2+)-induced Ca2+ release from the compartment. Two versions are used. In one, release is controlled by explicit thresholds, while in the other, release is a continuous function of cytosolic and compartmental [Ca2+]. Some model predictions are as follows. Starting with low Ca2+ influx and no spikes: (1) induction of spiking when Ca2+ influx is increased. Starting with spikes: (2) increase in magnitude and decrease in frequency when influx is reduced; (3) inhibition of spiking if influx is greatly reduced; (4) decrease in the root-mean-square value when influx is increased; and (5) elimination of spiking if influx is greatly increased. Since there is good evidence that hyperpolarizing spikes reflect cytosolic Ca2+ spikes, we used electrophysiological measurements to test the model. Each model prediction was confirmed by experiments in which Ca2+ influx was manipulated. However, the original spike activity tended to return within 5-30 min, indicating a cellular resetting process.     

Cloning of the aspartate-β-semialdehyde dehydrogenase structural gene fromEscherichia coli K12

Transformation ofEscherichia coli strains with the recombinant plasmid, prepared by shot gun cloning with pBR322 and containing the geneasd, the structural gene of aspartate--semialdehyde dehydrogenase, results in an increase in specific activity of 65-fold of the enzyme in crude extracts. Approximately 60 mg of pure enzyme may be obtained from 10 g of transformed cells (wet weight) in a simplified purification procedure. The molecular weight, amino acid composition, and kinetic properties of the enzyme appear to be the same as previously reported, and the first 36 amino acids of the N-terminal sequence have been determined.     

Electromyography of the gluteal muscles in Hylobates,Pongo, and pan: Implications for the evolution of hominid bipedality

The gluteal musculature of primates has been a focus of great research interest among those who study human evolution. Most current theorists agree that gluteus superficialis (= maximus) need not have changed its action in the step from pongid to hominid, but dispute has arisen over a purported change in action and role of the gluteus medius. To clarify the functions of gluteus medius, gluteus superficialis, and tensor fasciae femoris during ape locomotion, we conducted a telemetered electromyographic study of these muscles in two gibbons, one orangutan, and four chimpanzees as they walked bipedally on the ground and on a horizontal tree trunk, walked quadrupedally on the same substrates, and climbed a vertical tree trunk. The results indicate that the gluteus medius of apes is not, as has been previously suggested, primarily an extensor of the thigh; its action is chiefly that of medial rotation. The role of the gluteus medius during bipedality is the same in apes and humans–to provide side-to-side balance of the trunk at the hip. The change in the hominid lateral balance mechanism can be viewed as primarily osteological, allowing preservation of the same muscle function with an extended thigh. As a result, the stride length is increased and there occurs a diminution of the demands placed on other muscles to maintain anteroposterior balance at the hip and knee. Our data also support the view that vertical climbing may be specifically preadaptive to bipedalism. One may picture the earliest hominid as part biped, when on the ground traveling between scattered food trees, and part climber, when moving from the ground to food.     

Biofeedback treatment of dysmenorrhea

Nine dysmenorrheic women were run in EMG and thermal biofeedback procedures with concurrent autogenic relaxation practice. Significant reductions in subjective estimates of symptomology associated with dysmenorrhea were noted in all subjects. EMG levels correlated positively with the reductions in symptoms. Thermal levels did not correlate with EMG. In fact no consistent patterns in thermal measures were noted. However, thermal biofeedback cannot be ruled out as an effective treatment for dysmenorrhea since reductions in symptoms occurred during thermal biofeedback training. Another significant aspect of the present study is the effectiveness of long treatment procedures. A six month period was employed and significant reductions in symptoms were noted following two months of biofeedback treatment. Finally, the importance of beginning biofeedback treatment prior to onset of menstrual symptoms is indicated.     

The induction of chromosomal structural changes in male chickens by the alkylating agents: triethylene melamine and ethyl methanesulfonate

E. coli chromosomal DNA wastreated with various Pt co-ordiantion compounds and then used as donor DNA in E. coli transformation. Genetic analysis of transformants obtained with Pt-treated DNA showed effects of cis-diamminedichloroplatinum(II) (cis-Pt(II)) and cis-dimethyl-1,3-diaminopropane CL₄ (cis-Pt(IV) (DMDAP) on the processing of DNA. With trans-diamminedichloroplatinum(II) (trans-Pt(II)) appllied in similar concentrations no effects were found.The effects of cis-Pt(II) and cis-Pt(IV) (DMDAP) on the genetic processing were different. The effects of cis-Pt(II) could be explained by assuming intra-strand crosslinks as an important lesion.     

Competition by Estrogens for Catecholamine Receptor Binding In Vitro

Abstract: We have examined the ability of various steroids to compete for high-affinity binding of ³H-labeled ligands to catecholamine receptors in membranes prepared from rat cerebral cortex, striatum, and anterior pituitary. Ligands employed were: [³H]WB4101, [³H]prazosin, [³H]yohimbine, and [³H]clonidine (alpha-noradrenergic); [³H]dihydroalprenolol (beta-noradrenergic); [³H]spiperone and [³H]ADTN (dopaminergic). Only the 17β estrogens were effective and only binding of [³H]spiperone and [³H]ADTN in striatum and [³H]WB4101 and [³H]prazosin in cerebral cortex was reduced. Thus putative dopaminergic and alpha₁-noradrenergic sites alone appear to recognize estrogens. A slight competitive effect on [³H]spiperone binding to anterior pituitary membranes was also observed. Among the 17β estrogens tested, the most effective in all cases was the catechol estrogen 2-hydroxyestradiol (2-OHE₂). The ability of 2-OHE₂ (IC₅₀= 20–30 μM) to inhibit ligand binding to alpha₁ receptors was comparable to that of norepinephrine (IC₅₀= 10–20 μM), whereas for dopamine receptors in striatum and pituitary 2-OHE₂ was an order of magnitude less effective than dopamine (IC₃₀= 12 μM) in reducing binding of ³H ligands. Estradiol-17β and 2-hydroxyestrone were also able to inhibit binding, but the order of steroid potency was different for alpha₁ and dopaminergic receptors. Progesterone, testosterone, and corticosterone were without effect in all cases. These results show that there is specificity of steroid interactions with catecholamine receptors in the brain, both in terms of steroid structure and receptor type. The possible relevance of these interactions to neuroendocrine function is discussed.     

Intermediate Filaments of Schwann Cells

Abstract: Intermediate filaments were prepared from distal stumps of rabbit sciatic nerve 5 weeks after nerve section, at which time Schwann cells account for 85–90% of the cell area. A polypeptide of molecular weight 58,000 was the main component of this fraction. An antiserum raised in guinea pig against this polypeptide stained all cells present in the distal stump, as well as Schwann cells and 3T3 cells in culture. The identity of the molecular weight 58,000 polypeptide obtained from distal stumps with vimentin was proved with one and two-dimensional sodium dodecyl sulfate pol yacrylamide gel electrophoresis and with immunoautoradiography. It is concluded that the intermediate filament subunit of undifferentiated Schwann cells is vimentin. The possibility that Schwann cells in normal nerve may have another type of intermediate filament besides vimentin cannot be ruled out.     

Measurement of Methionine-Enkephalin [Arg6,Phe7] in Rat Brain by Specific Radioimmunoassay Directed at Methionine Sulphoxide Enkephalin[Arg6,Phe7]

Abstract: A specific and sensitive radioimmunoassay procedure for Metenkephalin[Arg⁶,Phe⁷] which allows its measurement in regions of the rat brain is described. The antiserum was raised against the methionine sulphoxide derivative of the peptide, and all samples and standards were oxidized with hydrogen peroxide prior to use in the assay with chloramine T-oxidized ¹²⁵I-labelled Met(O)-enkephalin[Arg⁶,Phe⁷]. The only significant cross-reactivity was 30% with the reduced heptapeptide Met-enkephalin[Arg⁶,Phe⁷]. The assay showed less than 0.15% cross-reactivity with fragments of the heptapeptide and with leucine-enkephalin-containing peptides. Acid acetone extraction of rat striatum followed by Sephadex G-50 chromatography and reverse-phase high pressure liquid chromatography showed that essentially all immunoreactivity co-chromatographed with Met-enkephalin[Arg⁶,Phe⁷]. This confirmed the specificity of the assay and showed that the striatum does not contain a high concentration of larger molecular weight forms with the heptapeptide at the COOH terminus. Distribution of the heptapeptide followed that of methionine enkephalin, with highest concentrations in the globus pallidus, intermediate levels in caudate-putamen and hypothalamus, and low levels in cortex and cerebellum.     

Antibodies Specific for α-N-Acetyl-β-Endorphins: Radioimmunoassays and Detection of Acetylated β-Endorphins in Pituitary Extracts

Abstract: Antibodies specific for α-N-acetyl-β-endorphins have been prepared by injecting into rabbits either α-N-acetyl-β-endorphin(1-31) or [α-N-acetyl, ε-acetyl-Lys⁹]-β-endorphin(1-9) linked by carbodiimide to bovine thyroglobulin. Both antisera were used to develop specific radioimmunoassays for α-N-acetyl-β-endorphins. The radioimmunoassays were used to measure α-N-acetylated β-endorphins in extracts of pituitary regions from different species. By comparison of the amounts of total β-endorphin and α-N-acetyl-β-endorphin immunoreactivity, a relative ratio of β-endorphin acetylation was obtained. The relative acetylation of β-endorphin was highest in rat posterior-intermediate lobe extracts (>90%). Beef and monkey intermediate lobes had a lower degree of acetylation (53 and 31%, respectively). Anterior lobe extracts from all three species contained low amounts of acetylated β-endorphin. Human pituitary extracts did not contain acetylated β-endorphins. By the use of cation exchange and high performance liquid chromatography, six different acetylated derivatives and fragments of β-endorphin were resolved in extracts of rat posterior-intermediate pituitaries. Two of these peptides corresponded to α-N-acetyl-β-endorphin(1-31) and -(1-27). One acetylated β-endorphin fragment had the same size as α-N-acetyl-β-endorphin(1-27) but was eluted earlier from the cation exchange column. This peptide had full cross-reactivity with antibodies directed against the middle and amino-terminal parts of β-endorphin. Compared with α-N-acetyl-β-endorphin(1-27), it had much less cross-reactivity with antibodies directed against the COOH-terminal part of β-endorphin, suggesting that it was a COOH-terminally modified derivative of β-endorphin(1-27). The remaining N-acetylated β-endorphin derivatives were eluted even earlier from the cation exchange column. The majority of these fragments were slightly larger in size than y-endorphin, i.e., β-endorphin(1-17), but smaller than β-endorphin(1-27). They had full cross-reactivity in an amino-terminally directed β-endorphin radioimmunoassay and a greatly diminished cross-reactivity with antibodies to the middle region of β-endorphin.