Biomedical subjects

R Einstein

Publications and source records attributed to R Einstein.

At least 19 recordsLinked to original sources

PubMed2002

Stressful animal housing conditions and their potential effect on sympathetic neurotransmission in mice.

Although the sympathetic nervous system (SNS) plays a major role in mediating the peripheral stress response, due consideration is not usually given to the effects of prolonged stress on the SNS. The present study examined changes in neurotransmission in the SNS after exposure of mice (BALB/c) to stressful housing conditions. Focal extracellular recording of excitatory junction currents (EJCs) was used as a relative measure of neurotransmitter release from different regions of large surface areas of the mouse vas deferens. Mice were either group housed (control), isolation housed (social deprivation), group housed in a room containing rats (rat odor stress), or isolation housed in a room containing rats (concurrent stress). Social deprivation and concurrent stressors induced an increase of 30 and 335% in EJC amplitude, respectively. The success rate of recording EJCs from sets of varicosities in the concurrent stressor group was greater compared with all other groups. The present study has shown that some common animal housing conditions act as stressors and induce significant changes in sympathetic neurotransmission.

Animals↗

PubMed2001

Apoptosis of ventricular and atrial myocytes from pacing-induced canine heart failure.

OBJECTIVE: Rapid ventricular pacing in dogs results in a low output cardiomyopathic state similar to idiopathic dilated cardiomyopathy in man. Cell death by apoptosis may play an important role in the loss of cardiac function. This study investigates the molecular pathways involved in the regulation of apoptosis in dogs with pacing-induced heart failure. METHODS: Apoptosis was identified by terminal transferase nick end-labelling (TUNEL) in the ventricles and atria of dog hearts affected by rapid-ventricular pacing. Western blots were used to determine expression of the components involved in the initiation (Fas, Fas-Ligand, FADD), regulation (Bcl-2, Bax) and execution (caspase-2 and caspase-3) of apoptosis. RESULTS: Pacing-induced heart failure resulted in a significant increase in the number of ventricular and atrial myocyte nuclei undergoing apoptosis as measured by TUNEL. Compared to the samples from control hearts (n=6) the expression of Bcl-2, an inhibitor of apoptosis, was significantly reduced in ventricles from five dogs with pacing-induced heart failure. No change in the expression of the apoptotic inducer Bax was detected. Fas and FADD were significantly elevated in all paced ventricles, and Fas-L was only detected in the paced hearts. Both caspase-2 and caspase-3 were elevated following ventricular pacing. CONCLUSIONS: We have identified components of the signalling pathways along which apoptosis proceeds following the induction of heart failure in dogs. Apoptosis was also detected in the atria raising the possibility that, like human dilated cardiomyopathy, the molecular changes are global.

Animals↗

PubMed2001

Sodium bicarbonate improves the chance of resuscitation after 10 minutes of cardiac arrest in dogs.

The likelihood of successful defibrillation and resuscitation decreases as the duration of cardiac arrest increases. Prolonged cardiac arrest is also associated with the development of acidosis. These experiments were designed to determine whether administration of sodium bicarbonate and/or adrenaline in combination with a brief period of cardiopulmonary resuscitation (CPR) prior to defibrillation would improve the outcome of prolonged cardiac arrest in dogs. Ventricular fibrillation (VF) was induced by a.c. shock in anaesthetised dogs. After 10 min of VF, animals received either immediate defibrillation (followed by treatment with bicarbonate or control) or immediate treatment with bicarbonate or saline (followed by defibrillation). Treatment with bicarbonate was associated with increased rates of restoration of spontaneous circulation. This was achieved with fewer shocks and in a shorter time. Coronary perfusion pressure was significantly higher in NaHCO3-treated animals than in control animals. There were smaller decreases in venous pH in NaHCO3-treated animals than in controls. The best outcome in this study was achieved when defibrillation was delayed for approximately 2 min, during which time NaHCO3 and adrenaline were administered with CPR. The results of the present study indicate that in prolonged arrests bicarbonate therapy and a period of perfusion prior to defibrillation may increase survival.

Animals↗

PubMed1999

Stress induced changes in transmitter release from sympathetic varicosities of the mouse vas deferens.

Activation of the sympathetic nervous system is an important component of the response to stress, but the effects of prolonged stress on sympathetic neurotransmission have not been assessed. In the present study we have examined the effect of 3 to 10 days exposure to stress induced by frequent handling and sham injections on neurotransmitter release from sympathetic varicosities of the mouse vas deferens. DiOC2(5)-fluorescence was used to visualise the sympathetic varicosities so that extracellular electrodes could be placed over known numbers of varicosities to monitor transmitter release using electrophysiological techniques. The frequency of excitatory junction currents (EJCs) increased with increasing duration of exposure to stress. The mean and maximum EJC amplitude significantly increased by 107% and 43%, respectively after 10 days of exposure to stress. The density of sympathetic varicosities innervating smooth muscle of the mouse vas deferens was not changed throughout the duration of the exposure to stress. The findings from this study demonstrate that the efficacy of transmitter release from the sympathetic varicosities is altered by repeated exposure of mice to stressful stimuli, such as handling and sham injections. Since such procedures are routine in many pharmacological experiments, it is important that investigators are aware of these changes so that due consideration is given when interpreting the data obtained from animals treated in this way.

Animals↗

PubMed1999

Changes in myocardial protein expression in pacing-induced canine heart failure.

Canine rapid ventricular pacing produces a low output cardiomyopathic state which is similar to dilated cardiomyopathy. In this study dogs were paced at 245 beats per minute (bpm) for 3-4 weeks until signs of heart failure were apparent. Unpaced dogs were used as controls. A previous study identified myocardial protein changes in the pH region 4-7 following ventricular pacing by using two-dimensional electrophoresis (2-DE) (Heinke et al., Electrophoresis 1998 19, 2021-2030). Many of these proteins were associated with mitochondria, energy metabolism within the cardiomyocyte, the cytoskeleton and calcium cycling. The present study aimed to examine the proteins migrating in the more basic region of the 2-DE pattern using immobilised pH gradient 3-10 strips to separate myocardial proteins. The expression of 31 proteins was altered in the paced myocardium: 21 were decreased and 10 increased. Following the identification of 23 of these spots by either amino acid compositional analysis or peptide mass fingerprinting or a combination of both, we confirm that many of the proteins whose expression is altered following ventricular pacing are associated with the mitochondria and energy production within the cardiomyocyte, including creatine kinase M, triosephosphate isomerase, phosphoglycerate mutase, cytochrome c oxidase, cytochrome b5, hydroxymethyl glutaryl CoA synthase, myoglobin, and 3,2-trans-enoyl-CoA transferase. Additionally, the cytoskeletal protein actin was increased in the paced hearts. These results strongly support the notion that energy production is impaired and mitochondrial dysfunction is involved in the development of heart failure in the paced dog.

Animals↗

PubMed1998

Protein changes observed in pacing-induced heart failure using two-dimensional electrophoresis.

Rapid ventricular pacing in dogs results in a low output cardiomyopathic state which is similar to idiopathic dilated cardiomyopathy in man. However, the pathophysiological mechanisms which cause this failure following pacing are unknown. Five dogs underwent rapid ventricular pacing. Hearts were stimulated at 245 beats per min (bpm) for four weeks and then reduced to 190 bpm to stabilize the failure. Six unoperated dogs were used as controls. This paper compares the two-dimensional gel electrophoresis (2-DE) protein patterns of left ventricular samples from the paced myocardium with the control dogs. Changes in protein expression were analyzed qualitatively and semi-quantitatively. In the paced dog samples 69 protein spots were significantly altered of which 42 were decreased and 27 were elevated. One qualitative change was observed: elongation factor Tu was present only the control hearts. Of these proteins, 20 have been identified by a combination of N-terminal protein microsequencing, peptide mass profiling by mass spectrometry, amino acid compositional analysis, and by comparison with databases of canine and human ventricular proteins. Ten of these are associated with mitochondria and energy production, including: pyruvate dehydrogenase E1 component, isocitrate dehydrogenase subunit alpha, HSP60 and HSP70, creatine kinase M and fatty acid binding protein. The cytoskeletal protein desmin was detected in reduced quantities and a spot corresponding to a fragment of desmin was increased. These results indicate that the development of heart failure in the paced dog involves alterations in mitochondrial energy production, the cytoskeleton and calcium activation.

Animals↗

PubMed1998

Ventricular fibrillation and defibrillation thresholds in sheep and dogs.

Ventricular defibrillation studies normally use dogs rather than other large species. To investigate the suitability of sheep, which are often cheaper and more readily available, we compared ventricular fibrillation and defibrillation thresholds (VFT, DFT) in sheep and dogs. A total of 12 sheep (31 +/- 5 kg) and six dogs (19 +/- 1 kg) were anesthetised with halothane. Fibrillation was induced via epicardial pacing leads, using a 1 s 50 Hz pulse. Biphasic defibrillation shocks were delivered across epicardial patches. Voltage-response curves for both fibrillation and defibrillation were generated. Logistic regression analysis was used to determine 50 and 90% probability of success for fibrillation induction and defibrillation. VFT was similar in sheep and dogs. DFT at 50% probability of success was significantly higher in sheep (369 +/- 14 V) than in dogs (299 +/- 31 V, P < 0.04) but within each species there was no correlation between heart weight and DFT. After defibrillation sheep took longer to return to sinus rhythm than dogs and electro-mechanical dissociation was observed in sheep, but not in dogs. Thus, sheep may not be an ideal model for ventricular defibrillation research but further studies of the intrinsic differences between sheep and dogs may provide insights into basic mechanisms of defibrillation.

Animals↗

PubMed1996

Type I IL-1 receptor: ligand specific confirmation differences and the role of glycosylation in ligand binding.

The extracellular domain of the type I Interleukin-1 receptor (sIL-1R) was expressed in Drosophila S2 cells as a secreted 43 kDa glycoprotein, as evidenced by its binding to Concanavalin A and enzymatic deglycosylation. sIL-1R bound IL-1 beta with a K(D) of 2 nM as determined by competition ELISA. N-Glycanase treated sIL-1R had a C. 100 fold lower affinity than glycosylated sIL-1R for IL-1 beta, suggesting that glycosylation is a key component of the IL-1 beta/IL-1 receptor interaction. Crosslinking of sIL-1R to (125)I-IL-1 beta could be competed with unlabelled IL-1 alpha, IL-1 beta, IL-1 receptor antagonist (IL-1ra), and a mutant of IL-1 (Th9Gly) which has reduced bioactivity but wild type receptor binding affinity. Limited proteolysis of sIL-1R in the presence of IL-1 alpha, IL-1 beta, IL-1ra, and Thr9Gly IL-1 beta with several different proteases followed by analysis of sIL-1R by Western blot was used to assess the effect of binding on sIL-1R conformation. While some proteases showed no differences in cleavage patterns or sensitivity between free and bound sIL-1R, others showed differences in either cleavage sites or sensitivity with different ligands. This implies that upon ligand binding there is a conformational change in the receptor which is sensitive to the particular ligand bound, and hence has implications for the ability of different ligands to trigger responses after binding to receptor.

Animals↗

PubMed1996

Changes in cardiovascular responsiveness to dopexamine and beta 1- and beta 2-adrenoceptor function after the chronic treatment of beta-adrenoceptor antagonists and agonists in anaesthetized dogs.

1. The studies were designed to investigate the changes in responsiveness to beta-adrenoceptor agonists induced by chronic administration of beta-adrenoceptor antagonists and agonists. 2. Dose-response curves for dopexamine, isoprenaline, noradrenaline and impromidine on heart rate, blood pressure and myocardial contractility (dP/dt:integrated isometric tension) were obtained in untreated dogs and compared to those measured in dogs which had been pretreated with propranolol (8 mg kg-1 day-1), atenolol (6 mg kg-1 day-1), isoprenaline (0.5 microgram kg-1 min-1) or noradrenaline (0.5 microgram kg-1 min-1) for 14 days. 3. Propranolol pretreatment significantly enhanced the inotropic and chronotropic responses to isoprenaline. There were noticeable, but non-significant increases in inotropic sensitivity to noradrenaline and dopexamine, especially at higher dose levels. In the atenolol treatment group, there were significant increases in inotropic responses to dopexamine and isoprenaline and in depressor responses to isoprenaline. 4. Thus, chronic administration of propranolol increased responses mediated by both beta 1- and beta 2-adrenoceptors, whereas, atenolol selectively enhanced the inotropic responsiveness to dopexamine, which is mediated mainly by beta 2-adrenoceptors. 5. Isoprenaline pretreatment caused a significant decrease in inotropic sensitivity to dopexamine, isoprenaline and noradrenaline and a significant reduction in chronotropic responses to dopexamine. The tendency to reduced depressor responses to isoprenaline and dopexamine failed to reach significance. Pretreatment with noradrenaline decreased only the inotropic response to noradrenaline. 6. Thus, chronic isoprenaline treatment reduced the responsiveness of both beta 1- and beta 2-adrenoceptors, while chronic noradrenaline infusion only reduced beta 1-adrenoceptor-mediated responses. 7. There was no significant change in any of the dose-response curves to impromidine after any beta-adrenoceptor antagonist or agonist treatment. This indicates that there was no non-specific alteration in responsiveness and that the observed changes were likely to be associated with specific alterations in beta-adrenoceptor number or function.

Adrenergic beta-Agonists↗

PubMed1996

Botulinum toxin A in non-dystonic tremors.

The present pilot study evaluated the effect of botulinum toxin A on primarily non-dystonic tremors using accelerometry in a single-blind, placebo-controlled design. Resting, postural, intention, or head tremor were assessed before and approximately 1 month after intramuscular saline and botulinum toxin A (25-50 U) respectively. Half of the patients showed > or = 30% placebo effect. Tremor in 10 of 17 patients (60%) studied improved further after botulinum toxin A (range 30-95%), exceeding the placebo effect by > or = 30%. Nine patients demonstrated clinically significant focal weakness in the extensor muscles after botulinum toxin A which interfered with fine movements. Patients were subdivided into PD-like and ET-like tremor(s). Both groups experienced large placebo effects for resting tremor, with little or no further improvement after botulinum toxin A. The improvement in postural tremor after botulinum toxin A, of 40% in the PD-like and 57% in the ET-like groups, however, was approximately twice that of placebo. In conclusion, botulinum toxin A exerts a modest tremorlytic effect, however the dose, and its distribution over the sites injected, need to be optimised to minimise focal weakness.

Adult↗

PubMed1995

Animal models of heart failure for pharmacological studies.

1. Animals for the study of congestive heart failure (HF) should have chronic, stable disease produced by methods which allow controllable damage and hence predictable disease severity. 2. Pressure and volume loading have commonly been used in the past but these methods are limited by the difficulty of controlling the disease severity. 3. HF induced by cardiotoxic agents, particularly doxorubicin, has been widely used for experimental purposes, but again, control of the degree of damage may be difficult. 4. Coronary artery ligation or occlusion produces heart failure in experimental animals, with clear clinical relevance. Disadvantages of such techniques include fatal arrhythmias and collateral vessel growth that prevents or slows the onset of HF. 5. A minimally invasive model of HF which is relatively controllable can be produced by repeated or single DC shocks across the left ventricle. 6. Chronic rapid ventricular pacing produces a technically simple, predictable, stable and controllable preparation of HF with neurohumoral and haemodynamic changes which mimic the clinical pattern. These changes are reversible in the short term but after pacing for 1 year or longer complete recovery does not occur after cessation of pacing. 7. It has recently been suggested that a single DC shock, three to seven times the threshold defibrillating current, administered to the left ventricle, might provide the basis for the development of a model of heart failure which is technically simple and controllable.

Animals↗

PubMed1995

'Atypical' tremor.

The tremor present in a small number of patients who present with features of both Parkinson's disease (PD) and essential tremor (ET) is ill-defined. We therefore studied 8 such patients with 'atypical' tremor and compared them clinically to 11 PD and 10 ET patients in a double-blind, cross-over, placebo-controlled trial. Tremor was assessed via accelerometry and surface EMG. Tremor frequency and the acute tremorlytic response to propranolol were not useful in discriminating between groups. Over 80% of atypical and PD patients showed alternating EMG, whereas 90% of ET patients had synchronous EMG activity. Tremor amplitude improved by > 90% in 38% of atypical, 27% of PD and none of the ET patients after single-dose L-dopa/benserazide (300/75 mg), a response which was significantly different between the ET and PD groups. Failure to differentiate electrophysiologically and pharmacologically between the atypical and other groups probably relates to clinical heterogeneity in the atypical group.

Benserazide↗

PubMed1995

Effect of enalapril in dogs with pacing-induced heart failure.

A repeated-measures study was conducted on 5 dogs to clinically, radiographically, and echocardiographically characterize the actions of the angiotensin-converting enzyme inhibitor, enalapril, before and after development of experimentally induced heart failure. Heart failure was artificially induced, using a surgically implanted programmable ventricular pacemaker, which stimulated the heart at a rate of 245 beats/min until a low-output cardiomyopathic state developed. This condition was then stabilized by decreasing the pacing rate to 190 beats/min. Pacing-induced heart failure was successfully induced in a mean +/- SD 4.2 +/- 1.95 weeks. The condition closely resembled the clinical, radiographic, and echocardiographic features of naturally acquired idiopathic dilated cardiomyopathy in dogs. Enalapril was well tolerated by dogs, and clinical adverse reactions did not develop. Results of echocardiographic studies indicated that enalapril treatment during the control period resulted in a significant (P < 0.05) increase in velocity of circumferential fiber shortening and a significant (P < 0.05) decrease in left ventricular ejection time. Therapeutic responses to enalapril were evident after development of heart failure. These included reduced severity of clinical signs of disease, evidence of decreased radiographically determined cardiac size (2 of 5 dogs), radiographic evidence of a reduction in pulmonary edema and congestion (4 of 5 dogs), significant (P < 0.05) reductions in left atrial and ventricular chamber dimensions (left atrial dimension, diastolic left ventricular internal dimension as determined echocardiographically), and improvement in some echocardiographic indices of left ventricular performance (velocity of circumferential fiber shortening and left ventricular ejection time).

Analysis of Variance↗

PubMed1994

Postural tremor of Parkinson's disease.

Previous studies have reported the resting tremor (RT) of Parkinson's disease to occur at frequencies between 3-7 Hz and to be characterised by an alternating pattern of electromyographic (EMG) bursting activity between opposing muscles. A postural tremor (PT), of higher frequency (> 6 Hz) and with a synchronous pattern of EMG activity, has also been previously described in Parkinson's disease. We investigated the electrophysiological and pharmacological properties of both the RT and PT of 11 patients with Parkinson's disease and 10 patients with essential tremor in a double-blind, placebo-controlled study of L-Dopa/benserazide and propranolol. Tremor amplitude and frequency were assessed via bidirectional accelerometry, and the pattern of activation of the antagonist muscles of the forearm was determined with use of surface EMG. In the Parkinson's disease group studied, the frequency, EMG pattern of bursts, and response to L-Dopa were similar for the two tremors (median improvement of RT by 70% and PT by 61%). Despite some overlap between the Parkinson's disease and essential tremor groups in the electrophysiology of the tremor, there was no such dramatic pharmacological response in the latter group. These results suggest that the RT and PT of Parkinson's disease share a common pathophysiology and are distinct from essential tremor.

Adult↗

PubMed1994

Cardiovascular actions of dopexamine in anaesthetized and conscious dogs.

1. Arterial blood pressure, heart rate and cardiac contractility were measured in pentobarbitone-anaesthetized mongrel dogs and in conscious, instrumented dogs. 2. In anaesthetized dogs (n = 5), dose-response curves were obtained by intravenous infusion of increasing doses of dopexamine (5-20 micrograms kg-1 min-1). Infusions were administered three times to each animal to determine whether the responses were reproducible. Dopexamine increased heart rate and myocardial contractility and decreased blood pressure. The dose-response curves for dopexamine did not differ significantly over time. 3. In a second group of dogs (n = 6), dose-response curves (5-20 mg kg-1 min-1) were obtained as above and repeated after the administration of amitriptyline (2 mg kg-1, i.v.). Amitriptyline caused a non-significant reduction in the inotropic and chronotropic responses to dopexamine. 4. Control dose-response curves for dopexamine (5-50 micrograms kg-1 min-1) were similarly obtained in a third group of dogs (n = 6), and repeated after bilateral vagotomy and sympathetic denervation of the heart. In these animals, a third dose-response curve for dopexamine was obtained after the administration of ICI 118551 (0.2 mg kg-1, followed by 0.2 mg kg-1 h-1). The chronotropic response to dopexamine was significantly reduced after cardiac denervation. There was a small, non-significant reduction in the inotropic and depressor responses after denervation. Administration of ICI 115881 significantly reduced both the inotropic and chronotropic response to dopexamine and caused a non-significant reduction in the depressor response. 5. The effect of raclopride (0.2 mumol kg-1, p.o.) was investigated by comparison of the dose-response curves for dopexamine in a control group of dogs (n = 6) to those obtained in dogs which had been pretreated with raclopride (n = 5). Raclopride had no significant effect on the cardiovascular responses to dopexamine. 6. In conscious, instrumented dogs (n = 5), pretreated with raclopride, dose-related positive inotropic and chronotropic and depressor responses to dopexamine infusions were recorded. The chronotropic responses in conscious animals were significantly greater than those in the anaesthetized animals.7. The results of this study indicate that both the positive inotropic and chronotropic actions of dopamine are due to a combination of direct, Beta2-adrenoceptor-mediated effects and the baroreceptor reflex response to the depressor action of the drug.

Adrenergic beta-Antagonists↗

PubMed1993

Mapping of neutralizing epitopes and the receptor binding site of human interleukin 1 beta.

Antibodies to synthetic peptides of human interleukin 1 beta (IL-1 beta) and to recombinant human IL-1 beta were used to identify epitopes of IL-1 beta associated with the neutralization of its biological activity. Analysis of antisera raised to 17 synthetic peptides derived from the mature IL-1 beta sequence showed that five regions (residues 6-15, 49-80, 58-80, 92-101, and 120-133) were both immunoprecipitating and neutralizing. Using a hexamer epitope mapping method, comparison of the regions recognized by four neutralizing rabbit antisera with those recognized by a rabbit antiserum raised to denatured IL-1 beta suggested two further neutralizing epitopes, residues 39-48 and 83-95. Finally, a neutralizing monoclonal antibody was shown to bind to the peptides 6-11 and 87-95 by peptide binding and mutagenesis. All of these regions appear predominantly on one face of IL-1 beta. The effect of mutations in residues 4-11 and 88-97, which lie within this face, on receptor binding and biological activity was determined. Most of the mutations tested affected both receptor binding and activity, whereas mutations in another face of IL-1 beta (residues 74-80) had no effect. Purification of two of the mutants with reduced bioactivity and receptor binding and analysis by two-dimensional NMR indicated no gross changes in tertiary structure. A third mutant had reduced bioactivity in two different bioassays but no change in receptor binding. Although two-dimensional NMR revealed no gross changes in conformation, small changes did occur at a site distal from that mutated. The data are consistent with other epitope mapping and receptor binding mutagenesis data and suggest that the neutralizing antibodies and receptor recognize different but overlapping regions of IL-1 beta.

Amino Acid Sequence↗

PubMed1992

Do changes in transcardiac impedance modulation correlate with haemodynamic status?

Implantable cardiac pacemakers and defibrillators have the ability to revert a variety of arrhythmias to normal sinus rhythm. For correct operation, such devices require accurate arrhythmia classification. Arrhythmia classification by these devices could be improved with the addition of a suitable haemodynamic sensor. This study investigated the use of transcardiac impedance for haemodynamic sensing. Ventricular fibrillation, ventricular tachycardia, electro-mechanical dissociation and five rates of ventricular pacing, each having a different associated level of haemodynamic compromise, were induced in each of seven mongrel dogs. The amplitude responses of the modulations of transcardiac impedance were compared with those of arterial pulse pressure (an established measure of haemodynamic status), and changes in cycle length. The correlation coefficient for changes in transcardiac impedance modulation amplitude and arterial pulse pressure was found to be 0.89. For transcardiac impedance modulation amplitude and cycle length, the correlation coefficient was 0.77, and for arterial pulse pressure and cycle length, the correlation coefficient was 0.85. In the acute anaesthetised dog, changes in the amplitude of transcardiac impedance modulations were shown to reflect different levels of haemodynamic status.

Animals↗

PubMed1991

Cell- and ligand-specific dephosphorylation of acid hydrolases: evidence that the mannose 6-phosphatase is controlled by compartmentalization.

Mouse L cells that possess the cation-independent mannose 6-phosphate (Man 6-P)/insulin-like growth factor (IGF) II receptor change the extent to which they dephosphorylate endocytosed acid hydrolases in response to serum (Einstein, R., and C. A. Gabel. 1989. J. Cell Biol. 109:1037-1046). To investigate the mechanism by which dephosphorylation competence is regulated, the dephosphorylation of individual acid hydrolases was studied in Man 6-P/IGF II receptor-positive and -deficient cell lines. 125I-labeled Man 6-P-containing acid hydrolases were proteolytically processed but remained phosphorylated when endocytosed by receptor-positive L cells maintained in the absence of serum; after the addition of serum, however, the cell-associated hydrolases were dephosphorylated. Individual hydrolases were dephosphorylated at distinct rates and to different extents. In contrast, the same hydrolases were dephosphorylated equally and completely after entry into Man 6-P/IGF II receptor-positive Chinese hamster ovary (CHO) cells. The dephosphorylation competence of Man 6-P/IGF II receptor-deficient mouse J774 cells was more limited. beta-Glucuronidase produced by these cells underwent a limited dephosphorylation in transit to lysosomes such that diphosphorylated oligosaccharides were converted to monophosphorylated species. The overall quantity of phosphorylated oligosaccharides associated with the enzyme, however, did not decrease within the lysosomal compartment. Likewise, beta-glucuronidase was not dephosphorylated when introduced into J774 cells via Fc receptor-mediated endocytosis. The CHO and J774 cell lysosomes, therefore, display opposite extremes with respect to their capacity to dephosphorylate acid hydrolases; within CHO cell lysosomes acid hydrolases are rapidly and efficiently dephosphorylated, but within J774 cell lysosomes the same acid hydrolases remain phosphorylated. This difference in processing indicates that lysosomes themselves exist in a dephosphorylation-competent and -incompetent state. Man 6-P-bearing acid hydrolases endocytosed by the L+ cells in the absence of serum were not distributed uniformly throughout the lysosomal compartment. The change in the dephosphorylation competence of L cells in response to serum suggests, therefore, that these cells contain multiple populations of lysosomes that differ with respect to their content of a mannose 6-phosphatase, and that serum factors affect the distribution of hydrolases between the different compartments.

Animals↗