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Biomedical subjects

R J Leonard

Publications and source records attributed to R J Leonard.

At least 55 records · Page 3Linked to original sources
PubMed1990

Differential effects of speech prostheses in glossectomized patients.

Five patients representing different categories of glossal resection were fitted with prostheses specifically designed to improve speech. Speech recordings made for subjects with and without their prostheses were subjected to a variety of analyses. Prosthetic influence on listeners' judgments of severity level/intelligibility, number of consonants in error, and on the acoustic measure F2 range of vowels was evaluated. Findings indicated that all subjects demonstrated improvement on the speech measures. However, the extent of improvement on each measure varied across speakers and resection categories. Implications of the findings for prosthetic speech rehabilitation in this population are discussed.

Adult↗
PubMed1990

An open-channel blocker interacts with adjacent turns of alpha-helices in the nicotinic acetylcholine receptor.

The binding site for an open-channel blocker, QX-222, at mouse muscle nicotinic acetylcholine receptors was probed using site-directed mutagenesis, oocyte expression, and electrophysiological analysis. The proposed cytoplasmic end of the M2 transmembrane helix is termed position 1'. At position 10' (alpha S252, beta T263, gamma A261, delta A266), Ala residues yield stronger and longer binding of QX-222 than Ser or Thr residues. These effects are opposite and roughly equal (30%-50% per mutation) to previously reported effects at position 6'. The polar end of an anesthetic molecule seems to bind to the position 6' OH groups, which provide a water-like region; the nonpolar moiety is near position 10' and binds more strongly in a nonpolar environment. Interactions with adjacent OH-rich turns of an amphiphilic helix may explain the widespread blocking effects of local anesthetics at the conduction pore of ion channels.

Amino Acid Sequence↗
PubMed1989

Developmental changes in the open time and conductance of acetylcholine receptors in aneural cultured Xenopus myocytes treated with cycloheximide or tunicamycin.

Aneural cultures of Xenopus myocytes were treated with cycloheximide or tunicamycin in order to determine the influence of continued protein synthesis and insertion of glycosylated acetylcholine (ACh) receptor proteins on developmental changes in ACh-activated single-channel events. The developmental stage of the cultures was estimated from the stage of intact embryos maintained at the same temperature as the cultures. Single-channel recordings (5 x 10(-7) M ACh) from untreated cultures revealed primarily low conductance (27.4 +/- 0.6 pS) events for the first 14 h in culture (temperature = 23-25 degrees C) and a second high conductance class of events (42.5 +/- 0.9 pS) at later developmental stages (after Stage 28). Treatment with cycloheximide (at Stage 28) or tunicamycin (at Stage 22) produced significantly (P less than 0.01) fewer high conductance events at later stages (Stages 31-47), but had no effect on the conductance or open time of the low conductance event. A significant decrease (Chi square, P less than 0.05) in ACh sensitivity was observed at Stage 27 in myocytes treated with tunicamycin at Stage 22. The results strongly suggest that the low and high conductance events represent two distinct receptor molecules synthesized at different developmental stages. The tunicamycin results also suggest that a developmental reduction in the mean open time of low conductance events in untreated cultures does not depend upon the continued insertion of newly synthesized receptors.

Acetylcholine↗
PubMed1989

Expression of Drosophila Shaker potassium channels in mammalian cells infected with recombinant vaccinia virus.

A recombinant vaccinia virus containing a Drosophila potassium channel (Shaker H4) cDNA was constructed by homologous recombination between wild-type vaccinia virus DNA and a transfer plasmid. The new virus was used to infect four types of mammalian cells in culture. Electrophysiological recording 24-72 hr after infection revealed the expression of voltage-gated transient potassium channels in all four cell types. The properties of the induced currents were identical to those previously observed following injection of the Shaker H4 transcript into oocytes. Vaccinia promises to be an effective vehicle for the heterologous expression of transmembrane ion channels in a variety of cell types.

4-Aminopyridine↗
PubMed1988

Evidence that the M2 membrane-spanning region lines the ion channel pore of the nicotinic receptor.

Site-directed mutagenesis and expression in Xenopus oocytes were used to study acetylcholine receptors in which serine residues (i) were replaced by alanines (alpha, delta subunits) or (ii) replaced a phenylalanine (beta subunit) at a postulated polar site within the M2 transmembrane helix. As the number of serines decreased, there were decreases in the residence time and consequently the equilibrium binding affinity of QX-222, a quaternary ammonium anesthetic derivative thought to bind within the open channel. Receptors with three serine-to-alanine mutations also displayed a selective decrease in outward single-channel currents. Both the direction of this rectification and the voltage dependence of QX-222 blockade suggest that the residues mutated are within the aqueous pore of the receptor and near its cytoplasmic (inner) surface.

Animals↗
PubMed1988

Vocal shadowing in singers and nonsingers.

Five nonsingers and five singers vocally matched the pitches of frequency modulated tones. Subject responses to the shadowing task were analyzed in terms of accuracy, and in terms of duration and speed characteristics of pitch changes. In addition, each frequency change was categorized according to hit, overshoot, undershoot, or oscillate patterns. Singers were found to effect pitch changes in significantly less time than the nonsingers. This finding was attributed to more direct patterns (hits) and faster maximum speeds obtained by the singers. Both groups demonstrated a direct relationship between the size of the pitch change and each of the duration and speed parameters examined. Similarly, for both groups, pitch lowering was faster than pitch raising. Faster speeds and more direct paths in effecting pitch changes were viewed as evidence of greater vocal proficiency in singers as compared to nonsingers. Implications of the data for mechanical and physiological aspects of voice frequency control are discussed.

Adult↗
PubMed1988

Early development of two types of nicotinic acetylcholine receptors.

Functional changes of acetylcholine receptor (AChR) channels in embryonic Xenopus myotomal muscle cells were examined during their development in culture. Single-channel currents evoked by 50 or 500 nM ACh were measured using the patch-clamp technique. In Xenopus myocytes the first emergence of AChRs takes place at about stage 20 (Nieuwkoop and Faber). Myotomes were dissociated at very early stages and plated in culture. Single-channel currents through AChRs were recorded at times ranging from a few hours (stage 21) to several days (stage 47) after the first emergence of AChRs. Two classes of AChR channel were recorded: One class had a low conductance with a long burst duration (low-conductance channel), and the other had a high conductance with a short burst duration (high-conductance channel). Both of these classes were active from the earliest time recorded (stages 21-24). One effect of development was a shift in the relative activity of the low- and high-conductance channels. Initially (stages 21-24), the low conductance channels predominated, accounting for over 95% of the observed events. After 3 d in culture, however, high- and low-conductance events occurred with roughly equal frequency. The other effect of development was a 4-fold decrease in the mean burst length of the low-conductance channel. The decrease in burst length took place rapidly, with about 60% of the change occurring within 24 hr in culture. The burst length of the high-conductance channel remained virtually constant during development, as did the unitary conductance of both channels and the voltage dependence of their mean burst lengths. The developmental change in the proportion of low- and high-conductance channels is likely due to the increased insertion of new high-conductance channels. However, the molecular mechanism of the shortening of burst length of the low-conductance channel is unknown.

Acetylcholine↗
PubMed1986

Acetylcholine raises excitability by inhibiting the fast transient potassium current in cultured hippocampal neurons.

The effects of acetylcholine on cultured hippocampal neurons were investigated by using the whole-cell version of the patch-clamp technique. The CA1 region of the hippocampus was excised from brain slices of young rats (12-19 day old), incubated in a papain solution, and dissociated. Neurons were plated on a glial feeder layer. The experiments were conducted mostly on neurons cultured for 2-6 days. Upon depolarization under voltage clamp, these cells exhibited a fast transient outward current (A-current), which was inhibited by 4-aminopyridine (2.5 mM). Acetylcholine (0.1 microM) also inhibited this A-current, as did the muscarinic agonists bethanechol and muscarine. As expected from their inhibition of the A-current, acetylcholine and 4-aminopyridine both increased the amplitude of the action potential and prolonged its duration. We conclude that the inhibition of the A-current constitutes a mechanism by which acetylcholine exerts its excitatory influence on hippocampal neurons.

4-Aminopyridine↗
PubMed1984

Differential development of two classes of acetylcholine receptors in Xenopus muscle in culture.

Physiological properties of acetylcholine receptors on muscle cells at very early stages of ontogeny were compared with those of cells at later stages. Two changes were observed that contributed to an overall shortening of the mean open time of single-channels. First, there was a shift in the relative proportions of two receptor types with different conductances and mean open times, such that the contribution of receptors with large conductance and short open time increased as development proceeded. Second, there was a sharp reduction in the mean open time of channels having small conductance, with no similar change in channels having large conductance.

Animals↗
PubMed1980

Antitumor activity of cytidine dialdehyde and its effects on nucleotide and nucleic acid synthesis.

Cytidine dialdehyde inhibited the growth of leukemia L1210 cells in culture at a 50% inhibitory concentration of 3.5 X 10(-5) M and, when administered i.p. at 200 mg/kg daily for 5 days, increased the mean survival of L1210 tumor-bearing mice by up to 171%. Given by the s.c. or i.v. routes, the compound was ineffective. The ethanol adduct of cytidine dialdehyde, although inactive in cell culture, increased the mean survival of L1210 tumor-bearing mice by up to 225% when administered i.p. but was inactive upon s.c. administration. Exposure of L1210 cells in culture for 25 hr to cytidine dialdehyde at the 50% inhibitory concentration increased the ribonucleoside di- and triphosphate pools, slightly increased deoxyadenosine triphosphate, deoxythymidine triphosphate, and deoxyguanosine triphosphate pools, and caused a pronounced increase in the deoxycytidine triphosphate pool. As determined by the rate of pyrimidine precursor incorporation into nucleic acids, this concentration of drug showed no effect on RNA synthesis but caused a reduction in DNA synthesis to 53% of control. Exposure of L1210 cells for 3 to 6 hr to 10(-4) M cytidine dialdehyde, a concentration which inhibits growth completely, effected an increase in the ribonucleoside di- and triphosphate pools and a rapid decrease of the deoxythymidine triphosphate pool. The deoxycytidine triphosphate and deoxyguanosine triphosphate pools decreased more slowly, and the deoxyadenosine triphosphate pool remained slightly elevated. Analysis of the rate of substrate incorporation into nucleic acids showed that this concentration of drug produced an 80% decrease in RNA synthesis and a 75% decrease in DNA synthesis 3 hr after drug exposure. These results suggest that the mechanism of action of cytidine dialdehyde may be due to its initial interference with DNA synthesis followed by a generalized inhibition of DNA, RNA, and protein synthesis at cytotoxic concentrations.

Animals↗
PubMed1979

Vocal shadowing under conditions of normal and altered laryngeal sensation.

Five adult male subjects vocally matched the pitches of tones in a steady-state tone matching task (SSTMT) and vocally shadowed frequency changes in a frequency modulated tone matching task (FMTMT). Matching was done in both a normal condition and a condition of topical anesthesia of the laryngeal mucosa. Pitch matching accuracy was not seriously altered in the anethesia condition, but anesthetized subjects required more time and had slower velocities in effecting transitions between tones on the FMTMT, as compared to normal. In both conditions there was a direct proportionality between interval size and each of the time and velocity variables examined, while frequency lowering was generally faster than frequency raising. Analysis of subject response strategies on the FMTMT revealed recurring patterns, described here as Overshoot, Undershoot, Oscillate and Hit. Fewer Hit, and more Oscillate, patterns were observed for the anesthesia condition as compared to normal. It was concluded that the FMTMT, as compared to the SSTMT, required finer and more rapid laryngeal control to coordinate respiratory driving pressures and glottal resistance, and hence was more susceptible to impairment of the laryngeal mucosal reflexogenic system. Implications of the data for modeling certain mechanical and physiological aspects of laryngeal control are discussed.

Adult↗
PubMed1976

Transplantation of the fetal rat pancreas: quantitative morphological analysis of islet tissue growth.

When pancreases from fetal rats were transplanted beneath the kidney capsule of isologous normal adult recipients, continued growth and differentiation of the endocrine portion of the pancreas occurred. While limited amounts of acinar tissues were identifiable in the early transplant period (7 days), such cells were absent in long term transplants (14 and 21 days). In contrast, while few definitive islet beta cells were present at the time of transplantation, following 21 days at the kidney site large circumscribed islets comprised of heavily granulated beta cells in association with duct epithelial cells predominated. Mitotic figures were seen in both these cell populations. Total islet mass had increased over 20-fold during the transplantation period. Similar results were observed if fetal pancreases were grown in organ culture for ten days prior to transplantation. Continued islet and duct cell growth, as evidenced by mitotic figures and an increase in absolute islet cell mass was obtained in such cultured explants when transplanted to either isogenic or allogenic recipients. These observations support the hypothesis that fetal pancreas may be the best source of donor material for transplantation to diabetic recipients, in part due to the continued growth and differentiation of the islet tissue during the transplantation period.

Animals↗
PubMed1976

Isotransplantation of organ-cultured neonatal pancreas: reversal of alloxan diabetes in the rat.

Pancreases from neonatal rats four to 16 days postpartum were grown in organ culture for from two to nine days. Approximately 10-20 explants, each measuring 1 mm.3 (1 mg.), were grown on a single Millipore filter placed at the gas-liquid interface of a medium consisting of 50 per cent horse serum and 50 per cent chick embryo extract. Following organ culture, an estimated 9-20 mg. of cultured islet tissue were dissociated with collagenase and isotransplanted into the peritoneal cavity of alloxan-diabetic recipients. In seven of eight recipients the diabetes was reversed between 11 and 53 days of posttransplantation. Animals receiving 12-16 mg. of cultured islet attained normoglycemia in 11-20 days; animals receiving 9-10 mg. of cultured islet tissue recovered between 45 and 53 days. These animals have remained symptom-free for over six months. Biopsies of grafts taken from the peritoneal cavity following reversal of diabetes contained well-vascularized islets compared primarily of heavily granulated beta cells. Quantitative analysis of host pancreases by the linear scanning method (biopsied at one to two weeks and four to five months following reversal of the diabetes) demonstrated that the total beta-cell mass was 3 per cent and the total insulin content was 6 per cent of the normal values. Little or no evidence of regeneration of host beta cells was observed. These studies show that a period of organ culture prior to isotransplantation does not impair the ability of islet tissue to reverse alloxan diabetes in the rat.

Animals↗
PubMed1976

Transplantation of islet tissue in the rat.

Long term reversal of alloxan diabetes has been accomplished by intraperitoneal isotransplantation of enzymatically dispersed neonatal pancreas. In contrast, allotransplanted recipients showed only a transient recovery from the alloxan diabetes followed by a return to the diabetic state at the time of the homograft rejection. These data strongly suggest that the reversal of the diabetic state was a consequence of the transplanted islets. This conclusion is further supported by quantitative analysis of biopsied pancreases from successfully reversed recipients which reveals only 3% of the normal beta cell mass. By comparison, recovery of transplanted islets composed primarily of aldehyde fuchsin positive beta cells was routinely accomplished in these recipients. Utilization of the more specific unlabeled immunoperoxidase method has revealed that some of the transplanted islets are composed of cells positive for glucagon and somatostatin, as well as insulin. Other recovered transplanted islets (generally smaller in size) are composed primarily of one cell type or the other. The presence of insulin, glucagon, somatostatin, and delete pancreatic polypeptide positive cells in the islets of normal rat pancreas has been confirmed. In addition, cells reacting positively for these hormones have been observed in the alloxan diabetic rat pancreatic islets and in islets from reversed recipients. The time required for the disappearance of glycosuria and hyperglycemia (usually occurring from one to eleven weeks posttransplantation) appeared to be related to the amount and age of the donor islet tissue transplanted. Fetal islet tissue was more effective on a per milligram basis in reversing the diabetic state. In addition, while reversal was obtained by transplantation of as little as 5 mg of neonatal islet tissue, relatively large amounts (20 mg) were required before successfully reversed recipients responded normally to glucose tolerance test. By comparison, a similar reversal of diabetes with normal response to glucose load was attained by transplanting only 3 mg of fetal islet tissue. Quantitative morphological evidence of large increases in absolute islet mass, obtained in fetal transplants at the renal subcapsular site suggests that the superiority of fetal islet donor tissue may by in its high growth potential. No adverse effects of an in vitro organ culture period, prior to transplantation, were observed with regard to the ability of neonatal tissue to reverse the diabetic state or for fetal islet tissue to continue to survive at the renal subcapsular site. Likewise, no advantage in regard to amelioration of the homograft rejection response was observed in cultured islet tissue; allotransplants of which were rejected at the kidney site.

Age Factors↗
PubMed1975

Plasma insulin and glucose levels in diabetic rats prior to and following islet transplantation.

Transplantation of islets of Langerhans into the peritoneal cavity of diabetic rats returns the recipients to a normal metabolic state and provides the capability to respond to a glucose stimulation with a rapid release of insulin. Of the two groups of transplanted diabetic rats studied, each had normal nonfasting glucose levels and normal glucose profiles following intravenous glucose tolerance tests. However, one group had significantly lower insulin levels than normal controls both under nonfasting conditions and 4 minutes following glucose stimulation. The other group demonstrated insulin levels greater than those of the normal controls when drawn under nonfasting conditions and essentially identical to the levels of the normal controls following a glucose load. Each transplanted group responded similarly to normal rats with a rapid release of insulin following glucose administration.

Animals↗