Large cell non-Hodgkin's lymphoma and Hodgkin's disease arising synchronously in a patient with chronic lymphocytic leukemia: importance of immunocytochemistry.
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Antigens, CD↗
Biomedical subjects
S M Schuetze
Publications and source records attributed to S M Schuetze.
Acquired protein S deficiency with multiple thrombotic complications after orthotopic liver transplant.
BACKGROUND: Orthotopic liver transplantation (OLT) is frequently complicated by thrombotic events that may threaten the viability of the allograft and severely compromise the overall outcome in these patients. Although multiple prothrombotic pathogenic mechanisms may be involved, a role for inadequate natural anticoagulant levels in the early postoperative period has been postulated. METHODS: We describe a case of a woman who suffered multiple thrombotic complications after a second OLT. Prospective assays of procoagulant and natural anticoagulant factor levels, in addition to screening tests for a variety of inherited and acquired hypercoagulable states, were carried out. RESULTS: Serial studies confirmed an acquired, isolated deficiency of Protein S associated with the second transplanted liver. Protein S levels were normal after the patient's first and third OLTs. There was no laboratory evidence of other underlying prothrombotic conditions. CONCLUSIONS: This unusual case of acquired Protein S deficiency demonstrates that the hypercoagulable phenotype may develop in the recipient of a liver from a heterozygous deficient donor. Furthermore, isolated low Protein S may be causally associated with hepatic artery thrombosis after OLT.
Adult↗
Substance P modulates single-channel properties of neuronal nicotinic acetylcholine receptors.
Substance P (SP) is present in avian sympathetic ganglia and accelerates the decay rate of acetylcholine (ACh)-evoked macroscopic currents in sympathetic neurons. We demonstrate here that SP modulates ACh-elicited single channels in a manner consistent with an enhancement of ACh receptor (AChR) desensitization. Furthermore, since AChR channel function was monitored in cell-attached patches with SP applied to the extra-patch membrane, the peptide must act via a second messenger mechanism. SP specifically decreases the net ACh-activated single-channel current across the patch membrane by decreasing both channel opening frequency and mean open time kinetics. These experiments demonstrate that a peptide can modulate neuronal AChR function by a second messenger mechanism.
Acetylcholine↗
Functional properties and developmental regulation of nicotinic acetylcholine receptors on embryonic chicken sympathetic neurons.
Measurement of acetylcholine (ACh)-induced currents indicates that the sensitivity of embryonic sympathetic neurons increases following innervation in vivo and in vitro. We have used single-channel recording to assess the contribution of changes in ACh receptor properties to this increase. Early in development (before synaptogenesis), we detect three classes of ACh-activated channels that differ in their conductance and kinetics. Molecular studies indicating a variety of neuronal receptor subunit clones suggest a similar diversity. Later in development (after innervation), changes in functional properties include increases in conductance and apparent mean open time, the addition of a new conductance class, as well as apparent clustering and segregation of channel types. These changes in channel function are compatible with the developmental increase in ACh sensitivity.
Acetylcholine↗
Embryonic acetylcholine receptors guarantee spontaneous contractions in rat developing muscle.
Many proteins are expressed in distinct embryonic and adult forms. However, in most cases we do not know why the embryonic form of proteins is required. This question can be readily addressed for the acetylcholine receptor (AChR) because developmentally specified modifications of this ligand-gated ion channel can be directly related to changes in membrane currents. In developing rat soleus muscle, spontaneous transmitter release causes miniature end-plate currents (m.e.p.cs) to flow into the muscle cell. We show here that these m.e.p.cs in neonatal soleus trigger spontaneous contractions. By injecting m.e.p.cs into young fibres, we showed that only embryonic m.e.p.cs can trigger such contractions; adult m.e.p.cs do not last long enough. Developing muscle fibres must be active for synapse and muscle differentiation. Our experiments indicate that the embryonic form of the AChR is essential for spontaneous contractile activity and may therefore be required for normal neuromuscular development.
Action Potentials↗
Kinetic differences between embryonic- and adult-type acetylcholine receptors in rat myotubes.
1. The burst structures of embryonic-type (low-gamma) and adult-type (high-gamma) nicotinic acetylcholine (ACh) receptors (AChRs) in rat myotubes were investigated with the patch clamp technique. The channels were activated with the agonists ACh and suberyldicholine (SubCh). 2. With either agonist, the distribution of burst durations showed two exponential components for both channel types: a 'long' component that corresponds to bursts of one or more openings and a 'brief' component that includes short, isolated openings. 3. For low-gamma channels, the percentage of all openings associated with the brief component decreased from approximately 40% at 10-100 nM-ACh to less than 10% at 10-100 microM-ACh. 4. Both high-gamma and low-gamma long bursts were interrupted by brief (30-90 microseconds) closures and longer (approximately 1 ms) transitions to a partially open (subconductance) state. The duration of brief closures and partial openings was relatively independent of the agonist, but their frequency within low-gamma bursts was 3-fold higher with SubCh than with ACh. 5. Brief closures are interpreted as transitions to a closed, doubly liganded state from which the channel can reopen. This model predicts that the channel opening rate is greater than 10,000 s-1 for both channel types at room temperature. 6. Estimates of the channel opening rate inferred from the rising phase of miniature end-plate currents recorded from rat soleus fibres are consistent with this interpretation. 7. Both high-gamma and low-gamma channels apparently operate via similar gating mechanisms. Differences in their gating behaviour can be explained in terms of faster kinetic rate constants for high-gamma channels.
Acetylcholine↗
Neural regulation of properties of the nicotinic acetylcholine receptor.
During nerve-muscle synapse formation, acetylcholine receptors become localized and modified to allow efficient transfer of information from nerve to muscle. In this paper we summarize our studies on two aspects of receptor modulation--their concentration at synaptic sites and their ability to desensitize in response to prolonged application of agonist. We demonstrate that receptor localization is a complex event which extensively reorganizes the structure of the junctional region. This allows the subsequent influences of contraction to be exerted differently in junctional and extrajunctional regions. We indicate that increases in muscle cell Ca2+ appear to mediate some of the effects of muscle contraction and suggest how regulation of Ca2+ levels may specify junctional and extrajunctional differences. Finally, we discuss the role of receptor phosphorylation in determining the rate of desensitization.
Acetylcholine↗
Desensitization of acetylcholine receptors in rat myotubes is enhanced by agents that elevate intracellular cAMP.
Incubating skeletal muscle fibers with forskolin, an activator of adenylate cyclase, increases the rate at which nicotinic acetylcholine receptors (AChRs) desensitize when exposed to ACh. Several reports indicate that this is due to the phosphorylation of AChRs by cAMP-dependent protein kinase, but other studies suggest that forskolin interacts with AChRs directly and that second-messenger systems are not required. To help clarify this issue, we studied the effects of forskolin and several other drugs on AChR function in embryonic rat myotubes. AChR function was studied by recording ACh-induced membrane depolarizations and ACh-induced single-channel currents. Our results indicate that forskolin at low concentrations enhances AChR desensitization through the action of a second messenger, most likely cAMP. An analog of forskolin that is much less effective in activating adenylate cyclase (1,9-dideoxyforskolin) is also much less potent in enhancing desensitization. Forskolin at low concentrations does not alter single-channel conductance or mean channel open time. However, when used at concentrations above 20 microM, forskolin may also exert direct drug effects on AChRs.
Animals↗
Development of rat soleus endplate membrane following denervation at birth.
Rat soleus endplates develop some of their characteristic features before birth and others after birth. Specializations appearing before birth include a localized cluster of acetylcholine receptors (AChRs), an accumulation of acetylcholinesterase (AChE) in the synaptic basal lamina, and a cluster of nuclei near the endplate membrane. In contrast, postsynaptic membrane folds are elaborated during the first three weeks after birth. We denervated soleus muscles on postnatal day 1, before folds had appeared, and followed the subsequent development of endplate regions with light and electron microscopy. We found that the denervated endplates initiated fold formation on schedule and maintained their accumulations of AChRs, AChE, and endplate nuclei. However, the endplates stopped fold formation prematurely and eventually lost their rudimentary folds. At about the same time, the junctional AChR clusters were joined by ectopic patches of AChRs. The former endplate regions also became unusually elongated, possibly as a consequence of the lack of membrane folds. Apparently, endplate membranes have only a limited capacity for further development in the absence of both the nerve and muscle activity.
Animals↗
Functional inhibition of acetylcholine receptors by antibodies in myasthenic sera.
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Acetylcholine↗
Developmental regulation of nicotinic acetylcholine receptors.
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Animals↗
Forskolin increases the rate of acetylcholine receptor desensitization at rat soleus endplates.
We have studied the function of acetylcholine (AcCho) receptors (AcChoRs) in rat soleus endplates before and after exposing the muscles to forskolin, a potent activator of adenylate cyclase. AcChoR function was tested by recording the membrane depolarization evoked by pulses of ionophoretically applied AcCho. Brief (2 msec) AcCho pulses delivered at 7 Hz evoked constant responses at untreated endplates. In contrast, after 10-100 microM forskolin was added to the bath, responses to similar pulse trains fell by as much as 80% within 1 sec. AcCho sensitivity recovered completely in less than 1 min after the pulses were stopped but fell again when the pulses were resumed. Similarly, longer (1 sec) ionophoretic AcCho pulses evoked roughly constant responses at control endplates, but after forskolin treatment the depolarization fell by one-half within less than 200 msec. These results indicate that forskolin increases the rate at which AcChoRs desensitize when exposed to agonist. Focal extracellular recordings showed that 20-100 microM forskolin also increased the decay rate of miniature endplate currents, indicating that forskolin may decrease AcChoR channel open time. Inhibitors of cAMP phosphodiesterase increased the potency of forskolin. When used alone, these inhibitors had effects similar to those of forskolin but smaller. Patch-clamp experiments indicated that forskolin at 100 microM may also interact with AcChoR channels directly, but at 20 microM this effect is negligible. Therefore, it is likely that the forskolin effects were mediated primarily by increased levels of intracellular cAMP.
Adenylyl Cyclases↗
Apparent acetylcholine receptor channel conversion at individual rat soleus end-plates in vitro.
Miniature end-plate currents (m.e.p.c.s.) were recorded extracellularly from individual fibres in neonatal rat soleus muscles for 2-24 h. In agreement with previous studies, the decay phases of m.e.p.c.s at many end-plates were doubly exponential with time constants of approximately 6 ms and approximately 1.5 ms at 21 degrees C. Earlier studies have shown that doubly exponential decays are due to the combined action of embryonic-type acetylcholine (ACh) receptors (AChRs) with long channel open times and adult-type AChRs with brief open times. When individual end-plates with doubly exponential m.e.p.c.s were studied for several hours or more, the relative size of the slow decay component frequently decreased with time. There was no evidence for a corresponding decrease in total m.e.p.c. amplitude. The time constants of the fast and slow components did not change. M.e.p.c. decays were stable at end-plates that were either very mature (small slow decay component) or very immature (small fast decay component). In these cases, the decay phases were virtually singly exponential and the time constant did not change. These data indicate that at end-plates with a mixture of adult-type and embryonic-type channels, the fraction of adult-type AChRs increases with time. This is similar to what occurs at end-plates developing in vivo. The results of ACh noise analysis experiments support this interpretation.
Action Potentials↗
Myasthenic serum selectively blocks acetylcholine receptors with long channel open times at developing rat endplates.
We have examined the physiological effects of antibodies from a highly specific myasthenic serum on acetylcholine receptors at developing rat endplates. The antibodies reduced the amplitude of miniature endplate potentials by 60% in 3- to 6-day-old animals but had no effect after day 14. Between days 7 and 12 the antibodies had an intermediate effect. This is the same period during which acetylcholine receptors with long channel open times (slow channels) disappear and receptors with short open times (fast channels) increase in number. Therefore, we examined the effect of the antibodies at endplates with a mixture of channel types more carefully. At all times tested, both noise analysis and analysis of miniature endplate currents showed that the antibodies reduced slow channel activity selectively. Single-channel recordings indicated that acetylcholine receptors that remained active after antibody treatment had normal gating properties. Thus, the antibodies appeared to silence slow channels selectively.
Animals↗
Gating properties of acetylcholine receptors at developing rat endplates.
The gating properties of acetylcholine receptors (AChRs) change during the development of rat soleus endplates. During the first 3 weeks after birth, the apparent mean channel open time (tau) decreases severalfold and the single-channel conductance (gamma) increases 50%. To better understand this phenomenon, we used a combination of noise analysis, analysis of miniature endplate currents (MEPCs), and single-channel recordings to quantify the relative levels of fast and slow AChR activity at developing soleus endplates. When the same endplates were studied with both noise analysis and MEPC analysis, results obtained with the two techniques were strongly correlated, but MEPC analysis yielded higher estimates of the relative amount of slow channel activity. Experiments designed to examine the distribution of fast and slow channels gave no evidence for a gradient of either channel type within individual endplates; rather, fast and slow channels appeared to be mixed together. However, the relative amount of fast and slow channel activity did vary markedly among different endplates within individual muscles. In agreement with earlier studies, we found a progressive decrease in the relative amount of slow channel activity during the first 3 weeks after birth. However, our data indicate that this process begins sooner than reported previously and takes longer to complete. Some of the same endplates that were studied physiologically were also examined in the electron microscope to test the hypothesis that changes in AChR gating might be related to ultrastructural changes such as the formation of folds. The physiological and ultrastructural results were essentially uncorrelated.
Age Factors↗
Transformation by Rous sarcoma virus prevents acetylcholine receptor clustering on cultured chicken muscle fibers.
Acetylcholine receptors aggregate in the membrane of cultured chicken myotubes; the process of receptor clustering can be stimulated by exogenous factors that we, among others, have begun to characterize. Chicken myoblasts transformed by temperature-sensitive mutants of Rous sarcoma virus, such as tsNY68, fuse to form multinucleated myotubes at 42 degrees C, the nonpermissive temperature for transformation. However, tsNY68-infected myotubes do not cluster acetylcholine receptors at 42 degrees C, even in the presence of active clustering agents. This phenomenon is not merely a result of viral infection, since myotubes infected with a transformation-deficient viral mutant, td107A, behave like noninfected myotubes with respect to receptor clustering; thus, the effects of tsNY68 on the clustering process must be mediated by the src gene product. These experiments may provide a method of identifying essential elements of acetylcholine receptor clusters.
Animals↗
Neonatal denervation inhibits the normal postnatal decrease in endplate channel open time.
The apparent mean channel open time (tau) of acetylcholine receptors (AChR) at skeletal muscle endplates decreases greater than 3-fold during development. In rat soleus muscles, the change occurs between postnatal days 8 and 18 as channels with long apparent open times (tau = 4.5 msec) disappear while channels with short apparent open times (tau = 1.5 msec) increase in number. We studied the role of innervation in this process by denervating neonatal soleus muscles prior to channel conversion. Tau at the denervated endplates was assayed at various times between days 8 and 18 by using fluctuation analysis. We found that early denervation blocked, or at least delayed, channel conversion. Unexpectedly, there was enhanced extrajunctional ACh sensitivity in the innervated muscles contralateral to the denervated ones. This observation allowed us to compare the apparent open times of junctional AChRs with those of extrajunctional AChRs 200 micron distant in the same innervated fibers. In developing muscles, tau at the extrajunctional sites decreased in parallel with tau at the endplates. Thus, neural regulation of AChR channel gating extends well beyond the endplate boundaries.
Acetylcholinesterase↗
Dye transfer between cells of the embryonic chick lens becomes less sensitive to CO2 treatment with development.
During the 3-h developmental stage 14 in the chick, intercellular transfer of iontophoresed fluorescent dyes becomes less sensitive to the lowering of intracellular pH by either CO2 or acetate ions. Up to developmental state 14, dye transfer between lens cells is reversibly blocked by exposure to 50% CO2. Beyond stage 14, dye transfer between these cells is no longer reversibly blocked by elevated pCO2. Electronic coupling is present throughout lens development and is not reversibly blocked by high pCO2 at any stage. The gap junctions joining the lens cells show morphological changes at developmental stage 14. Up to stage 14, all gap junctions observed between chick lens cells have connexon assemblies that appear condensed or crystalline following routine freeze-fracture microscopy. Beyond stage 14, chick lens cells express gap junctions with both the condensed assemblies and the dispersed assemblies characteristic of adult lens gap-junction structure.
Acetates↗