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Biomedical subjects

K F Miller

Publications and source records attributed to K F Miller.

At least 37 records · Page 2Linked to original sources

Starting to add worse: effects of learning to multiply on children's addition.

A major stumbling block in acquiring a new skill can be integrating it with old but related knowledge. Learning multiplication is a case in point, because it involves integrating new relations with previously acquired arithmetic knowledge (in particular, addition). Two studies explored developmental changes in the relations between single-digit addition and multiplication. In the first study, third-graders, fifth-graders, and adults performed simple addition or multiplication in mixed- and blocked-operations formats. Substantial interfering effects from related knowledge were found at all age levels, but were more pronounced for younger subjects. Thus in the early stages of learning multiplication, one consequence of learning a new operation is interference in performance of an earlier, related, but less recently studied skill. Consideration of error patterns supported the view that the problem of integrating operations is a prominent one even in the early stages of mastering multiplication. Patterns of errors were generally consistent across all age groups, and all groups were much more likely to give a correct multiplication response to an addition problem than the reverse. A second, longitudinal study confirmed this finding, showing evidence for impaired performance of addition over time within individual children (second-, third-, and fourth-graders) tested on simple addition and multiplication over a 5-month period. Analysis of reaction times for addition indicated that second-graders in advanced math classes and third-graders in regular math classes tended to slow down over the year in responses to addition problems. Fourth-graders, on the other hand, tended to increase their speed of addition over the course of the year. Multiplication showed a different pattern during this period, with no evidence for slowing among children who were able to perform this task. Disruption of previously learned knowledge in the course of acquiring new skills provides evidence that new knowledge and old knowledge are being integrated. This kind of non-monotonic development may provide an empirical method for determining the functional limits of a domain of knowledge.

Adult↗

Effect of a synthetic platelet activating factor on steroidogenesis of cultured porcine granulosa cells.

An embryo-derived platelet activating factor has been demonstrated to play an important role in reproduction. This report examined the effect of various doses of a synthetic platelet activating factor on the production of progesterone by porcine granulosa cells in culture. Granulosa cells aspirated from ovarian follicles of prepubertal gilts were grown for 24 hours in Dulbecco's Modified Eagles Media: Ham's F-12 with 5% fetal bovine serum and 1 micrograms/ml insulin. Cells were washed once in serum-free media and then cultured for an additional 48 hours with 0 to 5000 ng/ml of the platelet activating factor in media containing either 0.25% bovine serum albumin or 1% fetal bovine serum. Cells grown with fetal bovine serum produced 50% of the amount of progesterone that was produced in the absence of serum. Low doses of the platelet activating factor caused a slight decrease in progesterone production. Higher doses (greater than 500 ng/ml) in serum-free media caused a marked decrease in progesterone production. Serum had a protective effect at high doses of platelet activating factor which was probably mediated by enzymatic degradation of the platelet activating factor. In summary, platelet activating factor had no stimulatory effect on production of progesterone by porcine granulosa cells in culture.

Animals↗

Inhibin suppresses human chorionic gonadotropin secretion in term, but not first trimester, placenta.

Inhibin is produced by the placenta, with serum concentrations rising throughout pregnancy. In contrast, hCG serum concentrations peak in the first trimester and are 80-90% lower at term. This study was designed to determine the effect of inhibin on hCG secretion both early and late in gestation. Villus tissue from 3 term and 3 first trimester (8-10 week) placentas was maintained in an in vitro explant culture model for 5 days. Tissue from each placenta was incubated with control medium in 24 replicate wells for the first 72 h. During the final 48 h, 12 wells received control medium, and 12 wells received medium containing 1% rabbit antiserum raised against the alpha-subunit (residues 1-32) of the human inhibin peptide. The antiserum demonstrated biological activity by increasing serum FSH concentrations in an immature female rat bioassay. The relative increase in hCG secretion at the conclusion of days 4 and 5 in control and antiserum-treated groups for each first trimester and term placenta were compared to pretreatment hCG concentrations on day 3. The relative increases in hCG secretion of first trimester control groups on day 4 (mean +/- SD, 34 +/- 11%) and day 5 (63 +/- 23%) were compared to those in antiserum-treated groups on day 4 (39 +/- 13%) and day 5 (54 +/- 5%) and showed no significant difference between groups on either day. The same comparison in term cultures showed the relative increases in hCG secretion of control groups on day 4 (31 +/- 10%) and day 5 (64 +/- 50%) to be significantly lower than those in antiserum-treated groups on day 4 (100 +/- 41%) and day 5 (150 +/- 108%; P less than 0.001). These findings suggest that inhibin suppresses hCG secretion in term, but not first trimester, placentas.

Animals↗

Expression and performance in transgenic pigs.

Recent research clearly shows that fusion genes can be microinjected into a pronucleus of an ovum and integrate into the pig genome. Animals with such fusion genes are called 'transgenic'. The percentage of injected ova that developed into transgenic pigs varied among experiments from 0.31% to 1.73%. The percentage of transgenic pigs that expressed the fusion gene ranged from 17% to 100%. Eleven different regulatory sequences have been used for fusion genes transferred into pigs. Some of these regulatory sequences directed strong gene expression, but control over level of expression was inadequate. Other regulatory sequences directed weak expression, but imparted only brief spikes of induced expression. The predominant gene coding sequences transferred were for growth-related hormones. Elevation of growth hormone (GH) in expressing transgenic pigs enhanced plasma concentrations of insulin-like growth factor-I (IGF-I), insulin, and glucose, improved feed efficiency about 15%, and markedly reduced subcutaneous fat compared to nontransgenic siblings. Growth rate was enhanced in some transgenic GH pigs but not in others, possibly due to dietary limits. The 'over-expression' of GH was detrimental to the general health of most transgenic pigs. The most prevalent problems were lethargy, lameness, and gastric ulcers. Gilts that expressed foreign GH genes were anoestrous. Boars that expressed foreign GH genes lacked libido, but their semen was fertile when used by artificial insemination. Six different fusion genes have been transmitted from transgenic founders to progeny. Most of the transgenic pigs that produced progeny transmitted the fusion gene as an autosomal dominant trait to about half of their progeny. Regulatory sequences that will permit full control of gene expression must be developed before the full potential of gene transfer in pigs can be realized.

Animals↗

Genetic engineering of livestock.

Genetic engineering of livestock is expected to have a major effect on the agricultural industry. However, accurate assessment of the consequences of transgene expression is impossible without multigenerational studies. A systematic study of the beneficial and adverse consequences of long-term elevations in the plasma levels of bovine growth hormone (bGH) was conducted on two lines of transgenic pigs. Two successive generations of pigs expressing the bGH gene showed significant improvements in both daily weight gain and feed efficiency and exhibited changes in carcass composition that included a marked reduction in subcutaneous fat. However, long-term elevation of bGH was generally detrimental to health: the pigs had a high incidence of gastric ulcers, arthritis, cardiomegaly, dermatitis, and renal disease. The ability to produce pigs exhibiting only the beneficial, growth-promoting effects of growth hormone by a transgenic approach may require better control of transgene expression, a different genetic background, or a modified husbandry regimen.

Agriculture↗

Expression of human or bovine growth hormone gene with a mouse metallothionein-1 promoter in transgenic swine alters the secretion of porcine growth hormone and insulin-like growth factor-I.

Endocrine profiles were examined in swine that had integrated and expressed a fusion gene consisting of mouse metallothionein-1 (MT) promoter fused to either a human (h) or bovine (b) GH structural gene. Eleven of 18 pigs that had integrated MT-hGH and eight of nine pigs that had integrated MT-bGH expressed the genes. The level of expression varied widely among pigs (14-4551 micrograms/l for MT-hGH and 23-1578 micrograms/l for MT-bGH). The level of expression varied over time within each pig with no general pattern. Concentrations of porcine GH (pGH) were lower in MT-hGH pigs that expressed the gene than in non-expressors or in littermate controls. Insulin-like growth factor-I (IGF-I) concentrations increased with age in all pigs and were raised threefold in pigs expressing either the MT-hGH or MT-bGH genes. Measurement of the foreign GH in samples taken at 15-min intervals failed to reveal any short-term fluctuations in concentration. Administration of hGH releasing factor (GRF) to pigs expressing MT-bGH resulted in attenuated release of pGH compared with that of contemporary controls. Concentrations of bGH did not change after GRF injection. Human and bovine GH expressed in transgenic pigs appear to be biologically active in that they induce IGF-I and suppress endogenous pGH secretion. The failure to find short-term fluctuations and the lack of response to GRF injections are consistent with a non-pituitary and non-GRF regulatable site of production.

Animals↗

Ovarian response to injections of charcoal-extracted porcine follicular fluid and porcine follicle-stimulating hormone in gilts fed a progesterone agonist (altrenogest).

This experiment was conducted to compare the negative effects of charcoal-extracted porcine follicular fluid (pFF) and the positive effects of purified porcine follicle-stimulating hormone (pFSH) on growth of follicles and on plasma hormone concentrations. Twenty gilts were fed altrenogest for 18 days (20 mg.day-1.gilt-1) to suppress spontaneous growth of large follicles (greater than 6 mm in diameter). Gilts, assigned at random to receive pFF and pFSH administered in a 2 x 2 factorial arrangement, were injected 9 times at 8-h intervals starting 48 h before the last feeding of altrenogest and ending 8 h before slaughter (24 h after the last feeding of altrenogest). Blood was collected periodically through vena cava catheters. Treatment groups and mean number of medium follicles (3 to 6 mm in diameter)/gilt at necropsy were 1) 20 ml of charcoal-extracted porcine serum i.v. + 4 ml saline i.m., 30.8; 2) 20 ml of pFF i.v. + saline i.m., 0.2; 3) serum i.v. + 8 micrograms of pFSH (USDA-pFSH-B1)/kg BW in saline i.m., 59.0; and 4) pFF i.v. + pFSH in saline i.m., 36.2. Injections of pFF decreased (p less than 0.01) and injections of pFSH increased the number of medium follicles, and the interaction of pFF and pFSH was not significant. Plasma FSH decreased (p less than 0.01) during pFF treatment of saline-injected gilts at a rate of 0.29 ng.ml-1.h-1. During pFSH treatment, plasma FSH increased (p less than 0.05) at statistically identical rates of 0.33 and 0.32 ng.ml-1.h-1 in serum- and pFF-injected gilts.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

Absorption of compounds in medium by the oil covering microdrop cultures.

Microdrops of medium under either paraffin or silicone oil are commonly used for culture of early mammalian embryos. Radiolabeled estradiol, progesterone, and androstenedione in drops of medium under oil decreased by 51, 89, and 77%, respectively, after 24-hr incubation. Up to 14% of labeled estradiol moved to another drop of medium by passing through the oil. Several other substances tested (FSH, leucine, glucose, lactate, sodium ion, PGE2, PGF2 alpha) did not pass into the oil. Both paraffin and silicone oils can alter the composition of culture medium by absorbing and transferring certain types of compounds.

Absorption↗

Progress on gene transfer in farm animals.

Transgenic pigs and sheep have been produced by the microinjection of single-cell zygotes and two-cell ova with linear molecules of mouse metallothionein I (MT) promoter/regulator fused to either the human growth hormone (hGH) or bovine growth hormone (bGH) structural genes. The foreign genes integrated into the chromosomes of 3 of 111 lambs or fetuses and 31 of 341 pigs or fetuses examined. Immunoreactive hGH or bGH was present in the plasma of two transgenic lambs and 19 transgenic pigs. The hGH concentration in plasma varied greatly among pigs and was unrelated to the number of gene copies that had integrated. Rate of growth was not enhanced in any of the transgenic pigs in comparison to their littermate controls. However, bGH and hGH exerted definite biological effects in transgenic pigs as evidenced by significantly depressed backfat measurements, elevated levels of insulin-like growth factor (IGF-I), stimulation of mammary development (by hGH) and reduction in porcine growth hormone (pGH) to nondetectable levels in plasma. Five of six founder transgenic pigs transmitted the MT-hGH gene construct to one or more progeny. Three progeny of a boar that expressed hGH also expressed the foreign gene.

Animal Husbandry↗

A simple in vivo bioassay for inhibin-like activity using ovariectomized ewes.

Long term ovariectomized ewes were used in a bioassay for inhibin-like activity. The concentration of FSH 6 to 7 hr after injection of follicular fluid (a rich source of inhibin), as a percentage of pretreatment, regressed on the log of the dose had a slope of -26.0 +/- 7.6 (5 replications, mean +/- SD) and an index of precision of .32 +/- .04. This system was rapid, relatively easy and specific for in vivo inhibin-like activity. This bioassay was also used to determine the relative potency of an affinity-purified fraction of follicular fluid.

Animals↗

Immunoaffinity chromatography of bovine FSH using monoclonal antibodies.

Bovine FSH (bFSH) was used to immunize BALB/c mice. Spleen cells were fused to the SP 2/0 cell line to produce hybridomas that secreted monoclonal antibodies to bFSH. One of these antibodies (USDA-bFSH-MC28) was extensively characterized and found to be a gamma 1 with kappa light chains, having extremely low cross-reactivity with other bovine pituitary hormones and with ovine and porcine FSH. The dissociation constant as measured by Scatchard analysis was 4.3 nmol/l, and proved to be in a very useful range for affinity chromatography. In an essentially one-step immunoaffinity chromatography procedure, bFSH was easily isolated in a single chromatographic step from crude anterior pituitary homogenate with better yield and with the same purity as classical chromatographic techniques.

Animals↗

Effect of charcoal-extracted porcine follicular fluid and 17 beta-estradiol on follicular growth and plasma gonadotropins in gilts fed a progesterone agonist, altrenogest.

Thirty-four gilts in two experiments were fed altrenogest for 18 d to block spontaneous growth of ovulatory follicles after luteolysis. They were injected with estradiol or charcoal-extracted porcine follicular fluid (pFF) to determine 1) whether gonadotropin secretion could be depressed and 2) whether exposure to reduced levels of gonadotropins would result in decreased numbers of medium follicles (3 to 6 mm in diameter). Gilts in Exp. 1 received treatments in a 2 X 2 X 2 factorial arrangement starting 48 h before the last feeding of altrenogest. Corn oil or estradiol (2 micrograms/kg body weight), 5 ml of charcoal-extracted porcine serum (pS) or pFF were injected im four times at 8-h intervals and gilts were sacrificed 24 or 96 h after last feeding of altrenogest. In Exp. 2, gilts received one of four treatments consisting of 1) pS, injected iv nine times at 8-h intervals starting 48 h before the last feeding of altrenogest; 2) pFF, with injection protocol the same as for pS; 3) estradiol injected im three times and 4) four times at 8-h intervals starting 0 and 24 h, respectively, before the last feeding of altrenogest. Compared with pS or corn oil, estradiol increased (P less than .001) plasma estrogen and decreased (P less than .05) plasma luteinizing hormone (LH) without a significant effect on plasma follicle stimulating hormone (FSH). Estradiol, compared with corn oil, decreased (P less than .01) the number of medium follicles from 24.8 to 0/gilt and decreased (P less than .05) the weight of ovarian follicular fluid from 4.2 to 2.1 g/gilt at 72 h after the first injection. Five milliliters of pFF had no significant effect on plasma gonadotropins or number of medium follicles. However, 20 ml of pFF, compared with pS, decreased (P less than .05) plasma FSH from 45 ng/ml to 9 ng/ml 32 h after the first injection, had no effect on plasma LH, decreased (P less than .01) the number of medium follicles from 29.2 to 2.2/gilt and decreased (P less than .01) follicular fluid weight from 3.9 to 1.6 g/gilt by 72 h after the first injection. These results indicate that estradiol or a non-steroidal component of follicular origin can decrease secretion of gonadotropins and suppress recruitment of medium follicles in the pig.

Animals↗

Treatment of ovariectomized ewes with bovine follicular fluid decreases secretion of FSH without changing secretion of LH.

Ovariectomized ewes were injected with 0, 0.25, 0.5, 1.0 or 2.0 ml of charcoal-extracted bovine follicular fluid. Treating ewes with 2 ml of follicular fluid resulted in a decrease in circulating concentrations of FSH to 72.8% of the pretreatment value. With smaller doses of follicular fluid, the magnitude of the decrease was less. Concentrations of LH did not change significantly. Pretreatment of ovariectomized ewes with estradiol and/or progestogen did not alter the magnitude of the FSH decrease. This action of follicular fluid extract fits the effect of the non-steroidal substance known as inhibin or folliculostatin.

Journal Article↗

Seasonal LH profile in ovariectomized cattle.

Two experiments were conducted to examine seasonal changes in circulating LH concentrations in ovariectomized heifers. In experiment 1, four Holstein heifers were ovariectomized in April 1977 during middiestrus. Blood samples were collected daily for 30 days surrounding each equinox and solstice for one year to examine changes in plasma LH levels at the time of seasonal photoperiod changes. The LH concentrations were highest during the winter solstice period and lowest during the summer solstice period. In addition, samples taken at two-week intervals indicated a distinct LH profile with maximal LH concentrations during November-April and minimal concentrations during May-October. In experiment 2, eight Holstein heifers were ovariectomized in June-July, 1979 and given an estradiol or a control implant in October. A distinct LH profile for the interval extending from January, 1980 to February, 1981 was found in the heifers that were not treated with estradiol. Concentrations were maximal during December-April and minimal during May-November. The LH profile followed a similar pattern in the estradiol-treated heifers; however, the overall profile was at a higher level. These data indicate that underlying seasonal reproductive mechanisms are present in cattle even though the species ovulates and breeds throughout the year.

Journal Article↗

Inhibition and subsequent rebound of FSH secretion following treatment with bovine follicular fluid in the ewe.

Plasma concentrations of gonadotropins were examined after treatment of ewes with bovine follicular fluid (FF) in four experiments. Mean concentrations of FSH were significantly decreased by FF treatment. The FSH depression appeared to continue throughout the length of treatment when the duration of treatment was 2-4 days. However, in an experiment in which the treatment period was 8 days, mean concentrations of FSH initially declined and then returned to control levels during the last 4 days of treatment. In all experiments, a rebound in FSH concentrations was found 24-36 h after cessation of FF treatment. The magnitude of this rebound appeared to be proportional to the degree of FSH suppression during FF treatment.

Journal Article↗

Determination of the major urinary metabolite of flurazepam in man by high-performance liquid chromatography.

A high performance liquid chromatographic method for the determination of N-1-hydroxyethylflurazepam, the major urinary metabolite of flurazepam, in human urine is described. Urine specimens were incubated enzymatically to deconjugate N-1-hydroxyethylflurazepam glucuronide (metabolite) and were then extracted at pH 9.0 to extract the metabolite. The extracts were chromatographed on a microparticulate silica gel column using automatic sample injection, isocratic elution at ambient temperature and UV monitoring at 254 nm. The internal standard was 7 chloro-5(2'-chlorophenyl) 1,3-dihydro-1-2-dimethylaminoethyl-2H-1,4-benzodiazepine-2-one. The recovery from urine, in the 0.5-25.0 microgram/ml range, was 96.5 +/- 11.5% (S.D.), and the sensitivity limit was 0.5 microgram/ml. The method was found to be specific for N-1-hydroxyethylflurazepam in the presence of intact flurazepam and other possible urinary metabolites of flurazepam. The method was successfully applied to urine specimens collected from human subjects following the administration of 30-mg single oral doses of flurazepam dihydrochloride.

Adult↗