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D L Bennett

Publications and source records attributed to D L Bennett.

At least 37 records · Page 2Linked to original sources
PubMed1997

Nerve growth factor treatment increases brain-derived neurotrophic factor selectively in TrkA-expressing dorsal root ganglion cells and in their central terminations within the spinal cord.

Using immunocytochemistry and in situ hybridization, we have examined the expression of brain-derived neurotrophic factor (BDNF) and of neurotrophin receptors in dorsal root ganglion cells. In the adult rat, BDNF mRNA and protein were found mainly in the subpopulation of cells that express the nerve growth factor (NGF) receptor trkA and the neuropeptide calcitonin gene-related peptide (CGRP). NGF increased BDNF within the trkA/CGRP cells to the extent that almost 90% of trkA cells contained BDNF mRNA after intrathecal NGF treatment, and 80-90% of BDNF-expressing cells contained trkA. Non-trkA cells that expressed BDNF included some trkC cells and some small cells that labeled with the lectin Griffonia simplicifolia IB4, a marker for cells that do not express trks. However, very few trkB cells expressed either BDNF mRNA or protein, and NGF did not increase BDNF expression in non-trkA cells. BDNF protein was anterogradely transported both peripherally and centrally. The central transport resulted in BDNF immunoreactivity in CGRP containing terminal arbors in the dorsal horn of the spinal cord, and this immunoreactivity was increased by NGF treatment. Electron microscopic analysis revealed that the BDNF immunoreactivity was present in finely myelinated and unmyelinated axons and in axon terminals, where it was most concentrated over dense-cored vesicles. Our data do not support an autocrine or paracrine role for BDNF within normal dorsal root ganglia, but indicate that BDNF may act as an anterograde trophic messenger. NGF levels in the periphery could influence dorsal horn neurons via release of BDNF from primary afferents.

Afferent Pathways↗
PubMed1997

Chronic illness perception in adolescence: implications for the doctor-patient relationship.

OBJECTIVE: To describe the inter-relationship of chronic illness severity as perceived by adolescents with both psychosocial well-being and objective measures of illness severity. Additionally to compared the adolescents' perception of illness severity with how their physicians believe that the adolescents perceive their illness severity. METHODS: The psychological well-being of 48 adolescents with either cystic fibrosis (CF) or insulin-dependent diabetes mellitus (IDDM) was measured by four standardized questionnaires. The adolescents' perception of severity of illness was measured using an original instrument (PSCI), and this measure was compared to their physicians' estimates of how the adolescents perceived the severity of their illness and clinical illness. RESULTS: There were 24 patients in both the CF and IDDM groups. Both groups were found to function well psychosocially; although, there were more patients with low self image compared to normative values. Depression and low self image were associated with a greater adolescent perception of illness severity. For both chronic illness groups, physicians' assessment of assumed adolescent perception of disease severity correlated with clinical indices of disease severity and was higher than the perception of illness severity reported by the adolescents. For adolescents with CF, but not with IDDM, perception of severity of chronic illness correlated with clinical indices. CONCLUSIONS: For adolescents with chronic illness, their perception of illness severity is an important indicator of psychosocial well-being. Physicians do not accurately infer their patients' perception of illness severity.

Adaptation, Psychological↗
PubMed1996

Expression and function of ryanodine receptors in nonexcitable cells.

We have used reverse transcriptase-polymerase chain reaction to investigate the expression of ryanodine receptors in several excitable and nonexcitable cell types. Consistent with previous reports, we detected ryanodine receptor expression in brain, heart, and skeletal muscle. In addition, we detected ryanodine receptor expression in various other excitable cells including PC 12 and A7r5 cells. Several muscle cell lines (BC3H1, C2C12, L6, and Sol8) weakly expressed ryanodine receptor when undifferentiated but strongly expressed type 1 and type 3 ryanodine receptor isoforms when differentiated into a muscle phenotype. Only 2 (HeLa and LLC-PK1 cells) out of 11 nonexcitable cell types examined expressed ryanodine receptors. Expression of ryanodine receptors at the protein level in these cells was confirmed using [3H]ryanodine binding. We also investigated the function of ryanodine receptors in Ca2+ signaling in HeLa cells using single-cell Fura-2 imaging. Neither caffeine nor ryanodine caused a detectable elevation of cytoplasmic Ca2+ in single HeLa cells. However, ryanodine caused a significant decrease in the amplitude of Ca 2+ signals evoked by repetitive stimulation with ATP. These studies show that ryanodine receptors are expressed in some nonexcitable cell types and furthermore suggest that the ryanodine receptors may be involved in a subtle regulation of intracellular Ca2+ responses.

Animals↗
PubMed1996

trkA, CGRP and IB4 expression in retrogradely labelled cutaneous and visceral primary sensory neurones in the rat.

The pattern of trkA expression in relation to other neurochemical markers (CGRP and IB4) was investigated in primary sensory neurones innervating either the skin or bladder. Retrograde tracing using the fluorescent marker Fast Blue was performed followed by histochemistry. A greater proportion of visceral afferents compared with cutaneous afferents were trkA-immunoreactive (75% and 43%, respectively). CGRP expression correlated with trkA expression in that it was higher in visceral afferents than cutaneous afferents (69% and 51%, respectively). IB4 expression was negatively correlated with trkA expression, being lower in visceral afferents compared with cutaneous afferents (29% and 43%, respectively). The results emphasise the heterogeneity of trkA expression (and hence nerve growth factor, sensitivity) in afferents innervating different targets, and furthermore suggest that it is predominantly the CGRP-expressing population of primary afferents that express trkA.

Animals↗
PubMed1996

NGF but not NT-3 or BDNF prevents the A fiber sprouting into lamina II of the spinal cord that occurs following axotomy.

There is a stereotypical pattern of primary afferent terminations within the mature spinal cord; however, this pattern is not immutable. Peripheral axotomy causes A fibers to sprout into lamina II, a region from which they are normally excluded. We have investigated the role of neurotrophins in this response. Rats which had undergone sciatic axotomy were treated intrathecally with NGF, BDNF, or NT-3. A fibers were visualized using transganglionic labeling with cholera toxin B subunit; small fibers were visualized using CGRP immunostaining. NGF (12 microg/day for 2 weeks), but not NT-3 or BDNF, prevented both the axotomy-induced reduction in CGRP staining within lamina II and the sprouting of A fibers into this region. It is likely that the prevention of A fiber sprouting is a secondary consequence of NGF rescuing small fibers. This effect of NGF on dorsal horn sprouting has implications both for our understanding of the maintenance of CNS connectivity and for the treatment of neuropathic pain states.

Animals↗
PubMed1996

Accelerated schedule of hepatitis B vaccination in high-risk youth.

OBJECTIVE: To perform a feasibility and immunogenicity study of an accelerated schedule of hepatitis B immunization for high-risk youth. METHODOLOGY: High-risk adolescents attending a youth health centre and nearby youth refuges were immunized with Engerix-B recombinant vaccine, 20 micrograms intramuscularly, at 0, 2 and 6 weeks. Serology was performed prior to immunization and 3 months after the third dose. RESULTS: Forty-two subjects (27 female) aged 13-20 years entered the study. Two (4.8%), already hepatitis B virus (HBV) seropositive, were excluded. Thirty-six of 40 subjects had one or more risk factors for HBV. Participants were often elusive, needing multiple attempts to establish contact. Twenty (50%) of the 40 completed three immunizations and all 14 studied developed anti-hepatitis B surface titres of > 100 mlU/mL (geometric mean titre 630 mlU/mL, 95% confidence intervals 309-1290). CONCLUSIONS: High-risk youth can be immunized against hepatitis B successfully using an accelerated schedule, but compliance is difficult.

Adolescent↗
PubMed1996

Postnatal changes in the expression of the trkA high-affinity NGF receptor in primary sensory neurons.

In development approximately 70-80% of dorsal root ganglion (DRG) cells are dependent on nerve growth factor (NGF) for their survival, while in the adult only some 40% of DRG cells express the high-affinity NGF receptor, trkA. This discrepancy suggests that trkA expression, and therefore neurotrophin sensitivity, may alter as the animal matures. We have tested this possibility by counting the number of L4/5 DRG neurons showing immunoreactivity for trkA in rats from the day of birth to postnatal day 14. We also examined changes in p75 and IB4 labelling. On the day of birth, 71% of DRG cells were found to express trkA. However, this percentage gradually fell with age and reached adult levels at postnatal day 14. The expression of p75 did not parallel that of trkA, remaining relatively constant at between 45 and 50% of cells from birth to postnatal day 14. Over the same period there was a marked increase in the proportion of cells which bind the lectin IB4 from 9 (day of birth) to 40% (day 14). Since in the adult the IB4 population consists of small cells which mostly do not express trkA, this finding suggests that the postnatal down-regulation of trkA occurs in this population. Consistent with this suggestion are the results of double labelling for trkA and IB4, which confirmed that at times intermediate between birth and postnatal day 14 there was a high degree of coexpression between these markers (which is absent in the adult). This result also suggests that the down-regulation of trkA is unlikely to be directly responsible for the emerging IB4 binding.

Animals↗
PubMed1996

MR imaging of anterior cruciate ligament injury: independent value of primary and secondary signs.

OBJECTIVE: The purpose of this blinded study was to test the value of primary and secondary signs, independent of each other, for diagnosis of a tear of the anterior cruciate ligament (ACL) by means of MR imaging. MATERIALS AND METHODS: MR images of the knee in 74 patients who had the status of the ACL confirmed arthroscopically were blindly reviewed for status of the ACL three times: with primary signs masked, with secondary signs masked, and with no signs masked. On the basis of the observed signs at each session, the status of the ACL was predicted and the confidence in that prediction was noted. The MR imaging predictions were compared with the arthroscopic findings. The results were analyzed with receiver operating characteristic curves and stepwise discriminant analysis. RESULTS: We found no difference in diagnostic performance when only primary signs were used and when both primary and secondary signs were used. When using primary signs only, our observers performed significantly better than when using secondary signs alone. However, with secondary signs alone, our observers performed better than if left to chance for predicting ACL status. Of the secondary signs, our statistical analysis found bone contusion, anterior translation of the tibia, and an uncovered posterior horn of the lateral meniscus to be the most useful for diagnosis. CONCLUSION: Secondary signs do have value for deciding ACL status independent of primary signs. However, our observers performed much better when using primary signs instead of secondary signs. In the clinical setting, secondary signs do not help significantly in the diagnosis of ACL tears by means of MR imaging.

Anterior Cruciate Ligament↗
PubMed1995

Calcium signalling. Cracking ICRAC in the eye.

Capacitative Ca(2+) entry produces the very rapid light response in Drosophila photoreceptors; studies using this amenable system may help unravel the machinery and mechanisms that underlie this Ca(2+)-entry pathway.

Amino Acid Sequence↗
PubMed1995

Cost of providing pharmaceutical services to hospice patients.

The cost to a hospital pharmacy department of providing pharmaceutical services for a hospice program was studied. Drug acquisition costs were tabulated for all prescriptions dispensed to patients in the hospice program from April 1, 1992, to March 31, 1993. Direct time studies were conducted to determine the average personnel time involved in dispensing a prescription for a hospice patient. To determine the personnel time associated with auxiliary activities, self-reporting and work-sampling techniques were used. Indirect and delivery costs were also calculated. Relevant resource inputs were identified, measured, and valued by using both observation and pharmacy records. A total of 5640 hospice prescriptions were dispensed by the outpatient pharmacy; these represented 30% of all prescriptions dispensed. The average cost of dispensing a hospice prescription was $14.91 (total annual costs less drug acquisition costs). The total annual cost of providing pharmaceutical services to the hospice's patients was $196,607, and the total annual reimbursement received from the hospice program and self-paying patients was $155,623; therefore, costs exceeded revenues by $40,984. The cost to a pharmacy department of providing pharmaceutical services to patients in a hospice program substantially exceeded revenues. Greater efficiencies, a change in the pricing structure, or both may be necessary.

Contract Services↗
PubMed1995

Putative capacitative calcium entry channels: expression of Drosophila trp and evidence for the existence of vertebrate homologues.

Capacitative calcium entry is a major pathway through which intracellular calcium stores are refilled after stimulation. It has been suggested that the protein encoded by the transient receptor potential (trp) gene expressed in Drosophila photoreceptors may be homologous with capacitative calcium entry channels. Expression of the trp gene product in Xenopus oocytes led to significant increases in calcium entry only when the intracellular calcium stores were depleted. Previous investigations have found trp to be uniquely expressed in Drosophila photoreceptors, but PCR cloning shows that homologous proteins exist in Calliphora, mouse brain and Xenopus oocytes. It is thus possible that capacitative calcium entry in Xenopus oocytes is mediated by a homologue of trp.

Amino Acid Sequence↗
PubMed1995

The biological effects of endogenous nerve growth factor on adult sensory neurons revealed by a trkA-IgG fusion molecule.

Evidence suggests that nerve growth factor (NGF) may function as a mediator of some persistent pain states. We have used a synthetic protein, trkA-IgG, to sequester endogenous NGF and block the survival effects of NGF on cultured sensory neurons. We show that administration of trkA-IgG produces a sustained thermal and chemical hypoalgesia and leads to a downregulation of the sensory neuropeptide CGRP (calcitonin gene-related peptide) in treated sensory neurons. Acute administration of the molecule blocks the hyperalgesia that develops with carrageenan-induced inflammation. These data suggest that peripherally produced NGF normally acts to maintain the sensitivity of nociceptive sensory neurons and that, in some inflammatory states, an upregulation of NGF is responsible for alterations in pain-related behaviour. Antagonists of NGF may therefore be of clinical use in treating some chronic pain states.

Animals↗
PubMed1995

Patient satisfaction as an outcome indicator of pain management during labor and delivery.

Increased emphasis is being placed on using various indicators to assess the quality of health care services. The purpose of this study was to assess the use of patient satisfaction in evaluating the pain management of patients admitted for childbirth. Three hundred eighty-nine patients who were admitted to the labor and delivery unit were asked to evaluate the technical and interpersonal aspects of the pain management process. One hundred seventy-seven patients (45.5%) responded to the questionnaire. The results showed that the aspects that received the lowest level of satisfaction were: 1) the patient's role in the decision of preferred type of pain medication; and 2) the use of a different medication after informing the health care professional that the original medication was not relieving their pain. No relationship was found between satisfaction and pain. This indicated that satisfaction was based on the patient's reaction to aspects of the service experience, and was not dependent on the type or intensity of pain experienced.

Analgesia, Obstetrical↗
PubMed1994

Smoothly graded Ca2+ release from inositol 1,4,5-trisphosphate-sensitive Ca2+ stores.

Stimulation of cells with Ca(2+)-mobilizing hormones often leads to the generation of temporally and spatially complex changes in the intracellular Ca2+ ion concentration ([Ca2+]i). To understand the mechanisms regulating Ca2+ release from intracellular stores more clearly, we investigated the ability of histamine to release Ca2+ stores under different experimental conditions, using video imaging of single Fura-2-loaded HeLa cells. In Ca(2+)-free medium, stepwise increases in histamine concentration released an increasing proportion of the intracellular Ca2+ pool. This pattern of Ca2+ release is analogous to the "quantal" release of Ca2+ previously observed using permeabilized cells. Quantal Ca2+ release was observed at both 20 and 37 degrees C and was not due to inactivation or desensitization of the Ca2+ release mechanism, since application of histamine in a pulsatile manner, which avoided desensitization of the Ca(2+)-release mechanism, still produced a quantal response. In Ca(2+)-containing medium at both 20 and 37 degrees C, stepwise increases in histamine concentration evoked [Ca2+]i responses where the amplitude was smoothly graded in direct proportion to the histamine concentration. Similar smoothly graded responses were observed from HeLa cells in Ca(2+)-free medium. These data indicate that hormone-evoked Ca2+ release from intracellular stores is limited by the hormone concentration, and that the mechanisms underlying complex [Ca2+]i signals do not lead to an all-or-none release of Ca2+ from the entire intracellular Ca2+ pool. We suggest that the hormone-sensitive intracellular Ca2+ pool is composed of functionally discrete units that are recruited by agonists in a concentration-dependent manner.

Calcium↗
PubMed1994

Postsynaptic effects of long-range afferents in distant segments caudal to their entry point in rat spinal cord under the influence of picrotoxin or strychnine.

1. Previous work has shown that substantial numbers of arriving myelinated afferent nerve fibers travel for many segments caudal to their entry point and terminate in the gray matter of distant segments. This fact is surprising because no monosynaptic post-synaptic responses attributable to these long-range afferents are observable in the distant segments. Evidence has been produced to explain this paradox by showing that impulse transmission is normally blocked in these long-range afferents by the tonic operation of a primary afferent depolarization (PAD) mechanism. Impulse transmission is restored if the PAD mechanism is disabled with gamma-aminobutyric acid antagonists. In this paper we examine the postsynaptic consequence of restoring conduction in the long-range afferents with picrotoxin. Because picrotoxin blocks PAD and increases the excitability of cells, we here contrast its action with that of the glycine antagonist strychnine, which increases the excitability of dorsal horn cells but does not affect PAD. 2. The preparation used throughout these experiments was decerebrate and spinal at T11. On one side dorsal roots T12, T13, and L1 were intact but all more caudal roots on that side were cut. Recordings of single units were made in the L6 segment with a rigid search pattern. The innervated area of skin on the flank was repeatedly stimulated with pressure in a repeated pattern. In the control state, few cells were detected responding in the L6 segment, which was five to seven segments caudal to the intact dorsal roots. After picrotoxin, the number of cells with excitatory receptive fields rose by a factor of 8, whereas there was no change in the number of inhibited cells. By contrast, after strychnine there was no change in the number of excited cells but the number of inhibited cells rose by a factor of 10. Details of the size of receptive fields, adequate stimuli, and location of the responding cells are provided. 3. To determine the number of synapses involved in producing the excitatory responses of L6 cells, we applied electrical stimuli to the peripheral receptive fields and to the L1 dorsal root. In the presence of picrotoxin, many cells were recorded with a latency consistent with monosynaptic connection. Some of these cells responded twice to two stimuli separated by 2 ms, but there was a latency variation between successive responses. Some cells responded with a short fixed latency and twice in 2 ms. No such cells were detected in the control state or in the presence of strychnine.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1992

The post-translational processing and intracellular sorting of PC2 in the islets of Langerhans.

Proinsulin conversion in the insulin secretory granule is mediated by two sequence-specific endoproteases related to the Kex2 homologues, PC2 and PC3 (Bennett, D. L., Bailyes, E. M., Nielsen, E., Guest, P. C., Rutherford, N. G., Arden, S. D., and Hutton, J. C. (1992) J. Biol. Chem. 267, 15229-15236; Bailyes, E. M., Bennett, D. L., and Hutton, J. C. (1992) Enzyme, in press). Radiolabeling studies using isolated rat islets showed that PC2 was synthesized initially as a 76-kDa glycoprotein which was converted by limited proteolysis to the mature 64-66-kDa form. Conversion was initiated approximately 1 h after synthesis and proceeded via intermediates of 71, 68, and 66 kDa with a t1/2 of 140 min. Release of only the 66- and 64-66-kDa radiolabeled forms of PC2 was induced by glucose and then only at times more than 2 h following synthesis. Proinsulin conversion, by contrast, was more rapid (delay = 30 min, t1/2 = 60 min), and release commenced as soon as 1 h after synthesis with the secreted material being comprised of the precursor, intermediate, and mature forms of insulin. Ultrastructural analysis of islet beta cells showed that PC2 was concentrated in secretory granules. Subcellular fractionation combined with immunoblot analysis showed that insulinoma secretory granules contained only the mature 64-66-kDa form of PC2, whereas fractions enriched in Golgi and endoplasmic reticulum contained a mixture of the 76- and 66-kDa forms of the enzyme. These results indicate that post-translational proteolysis of PC2 is initiated before sorting into the regulated pathway of secretion and that the relative proportions of proinsulin and PC2 packaged into secretory granules will change with physiological conditions.

Amidohydrolases↗
PubMed1992

Identification of the type 2 proinsulin processing endopeptidase as PC2, a member of the eukaryote subtilisin family.

Enzymological studies have implicated two Ca(2+)-dependent endopeptidases in the conversion of proinsulin to insulin; a type 1 activity which cleaves on the C-terminal side of Arg31-Arg32 and a type 2 activity which cleaves C-terminally to Lys64-Arg65 in the proinsulin sequence. The possibility that these enzymes are related to the recently discovered family of mammalian subtilisin-like gene products (furin, PC2, and PC3) and the yeast propheromone-converting enzyme (KEX-2), was investigated. Degenerate oligonucleotide primers flanking the putative catalytic domain within this gene family were used in a polymerase chain reaction to amplify related sequences from rat insulinoma cDNA. One major product of 700 base pairs was obtained which was greater than 99% identical to the corresponding rat PC2 sequence. This cDNA was subcloned into the bacterial expression vector pGEX-3X to generate a recombinant protein for antibody production. Western blot analysis showed the immunoreactivity was prominent in neuroendocrine tissues as a 65-kDa protein. It was concentrated in secretory granule-enriched fractions of insulinoma tissue, where it was present as a readily solubilized monomeric protein. Deglycosylation studies using endoglycosidase H and N-glycanase showed that the 65-kDa protein was comprised of approximately 9% carbohydrate, consistent with the presence of three consensus sequences for N-linked glycosylation in rat PC2. The immunoreactivity co-eluted with the type 2 proinsulin endopeptidase on gel filtration and ion-exchange chromatography and the antisera specifically immunoprecipitated type 2 activity from insulin granule extracts. N-terminal sequence analysis of the immunoreactive protein gave two sequences which corresponded to residues 109-112 and 112-119 of rat PC2. This indicated that posttranslational processing of PC2 itself occurs C-terminally to basic amino acids to produce the mature enzyme. It is concluded that PC2 is the type 2 endopeptidase involved in proinsulin conversion. Localization of PC2 immunoreactivity to other tissues of the diffuse neuroendocrine system suggests that the type 2 endopeptidase also functions in the processing of precursor forms of other prohormones and polypeptide neurotransmitters.

Amino Acid Sequence↗