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Psychiatric illness is associated with both chronic pain syndromes and opioid use disorder, further complicating how we care for patients with psychiatric issues. We report a unique case of a de novo and persistent psychotic disorder after complicated opioid withdrawal in a patient without any psychiatric history. The patient developed persistent auditory hallucinations after discontinuation of chronic opioid therapy that responded only to atypical antipsychotic (olanzapine) treatment. This case illustrates the neuropsychiatric effects of chronic opioid exposure, as well as layered clinical management dimensions related to opioid detoxification and psychosis treatment. Long-term opioid therapy may have lasting neuropsychiatric effects, including playing a role in the development and/or expression of psychotic disorders. Here we review the limited literature on the effects of opioids on psychosis. This complex case also demonstrates a clinical approach for effectively co-managing psychiatric symptoms in the context of chronic pain and chronic opioid therapy., Competing Interests: The authors declare no conflicts of interest., (Copyright © 2022 Wolters Kluwer Health, Inc. All rights reserved.)

Background: Given that training is integral to providing constructive peer feedback, we examined the impact of a regularly reinforced, structured peer assessment method on student-reported feedback abilities throughout a two-year preclinical Communication Skills course., Methods: Three consecutive 32-student medical school classes were introduced to the Observation-Reaction-Feedback method for providing verbal assessment during Year 1 Communication Skills orientation. In biweekly small-group sessions, students received worksheets reiterating the method and practiced giving verbal feedback to peers. Periodic questionnaires evaluated student perceptions of feedback delivery and the Observation-Reaction-Feedback method., Results: Biweekly reinforcement of the Observation-Reaction-Feedback method encouraged its uptake, which correlated with reports of more constructive, specific feedback. Compared to non-users, students who used the method noted greater improvement in comfort with assessing peers in Year 1 and continued growth of feedback abilities in Year 2. Comfort with providing modifying feedback and verbal feedback increased over the two-year course, while comfort with providing reinforcing feedback and written feedback remained similarly high. Concurrently, student preference for feedback anonymity decreased., Conclusions: Regular reinforcement of a peer assessment framework can increase student usage of the method, which promotes the expansion of self-reported peer feedback skills over time. These findings support investigation of analogous strategies in other medical education settings., Supplementary Information: The online version contains supplementary material available at 10.1007/s40670-021-01242-w., Competing Interests: Conflicts of interestOn behalf of all authors, the corresponding author states that there is no conflict of interest., (© International Association of Medical Science Educators 2021.)

An assessment of blood ammonia concentration is common medical practice in the evaluation of an individual with an unexplained mental status change or coma. The determination of a blood ammonia level is most commonly done using a glutamate dehydrogenase (GLDH)-based assay, although there are many potential sources of artifact and the literature is inconsistent regarding key preanalytic issues. Using a GLDH-based assay, we first investigated matrix effects using three anticoagulants: heparin, EDTA and oxalate. Heparin-anticoagulated plasma was substantially less precise than EDTA- and oxalate-anticoagulated plasma. Oxalate-anticoagulated plasma showed a greater baseline of apparent ammonia than either heparin- or EDTA-derived plasma, presumably due to interferants. We then evaluated the stability of EDTA-anticoagulated plasma for assessment of ammonia when stored at 4°C,-14°C or -70°C. There was a linear increase of ammonia with storage at both 4°C and -14°C. Plasma kept at -70°C for up to three weeks showed no change in measured ammonia relative to the baseline determination. This work clarifies preanalytic conditions for which a precise determination of ammonia can be accomplished using a GLDH-based assay., (Copyright © 2017 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.)

Our simple kinetic model, based on the classic "binding change mechanism", describes the stepping kinetics for the rotary enzyme motors. The model shows that the cooperative interactions between active sites in the motor enzyme F1-ATPase induce the stepping product release. This phenomenon results from non-harmonic oscillations in the enzyme forms. The found rate constants, corresponding to the stepping phenomenon, are close to the rate constants known for the F1-ATPase. The duration of dwells during the product release is shown to depend on the ATP concentration in accordance with the known experimental data., (Copyright © 2010 Elsevier Ireland Ltd. All rights reserved.)

A graphical analysis demonstrates the ability of slow substrate activation and certain types of cooperativity between the two enzyme active sites to generate sustained oscillations. The analysis allows us to estimate kinetic parameter values for which oscillations exist. The scheme analyzed can explain the cyclical changes in functioning of various motor enzymes. Moreover, this scheme does not generate bistability for any parameter values. The graphical analysis presented is simple and visually clarifies the regulatory role of the details in the kinetic schemes.

A graph-theoretic approach is shown to simplify the analysis of transient enzyme kinetics. The coefficients of the characteristic polynomial for kinetic equations are obtained by graphical construction of directed trees and sub-trees in the kinetic scheme. An example of kinetic schemes, providing a simple time-dependent analytical solution, is demonstrated. This example describes a substrate-inhibited enzymatic reaction and interprets the pH-dependent inhibition of the lactate dehydrogenase. It is shown that rapid equilibrium in some parts of the kinetic scheme can simplify the analysis. The enzyme pre-incubation with a product is shown to be characterized by the non-monotonous transient kinetics. This phenomenon is useful to estimate correctly the kinetic parameters. It is supposed that the lactate dehydrogenase substrate inhibition can be important for switching the glycolytic fluxes.

A kinetic model for dynein, a molecular motor, is considered. This model explains the oscillatory behaviour, observed by Chikako Shingyoji et al. [Ch. Shingyoji, H. Higuchi, M. Yoshimura, E. Katayama, T. Yanagida, Dynein arms are oscillatory force generators, Nature 393 (1998) 711-714.] and by Susumu Aoyama and Ritsu Kamiya [S. Aoyama, R. Kamiya, Cyclical interactions between two outer doublet microtubules in split flagellar axonemes, Biophys. J. 89 (2005) 3261-3268.] in surprisingly simple axonemal fragments. The model shows that sustained oscillations can be generated due to the obligate cooperative interaction of the two dynein heads in the axonemal fragments. No other feedback control interactions are involved in the model to explain oscillations, similar to those observed experimentally, for realistic dynein rate constants. The modified model shows how the ATP hydrolytic exhaustion influences the amplitude and frequency of dynein oscillatory activity.

Butyrylcholinesterase (BChE) displays hysteretic behavior with certain neutral and charged substrates in the approach to steady state. Previous studies led us to interpret this phenomenon in terms of slow transitions between two enzyme conformers E and E'. This kinetic peculiarity is observed in human, horse and rat BChE. Oscillations that superimpose on the hysteretic lag are observed when benzoylcholine and N-alkyl derivatives of benzoylcholine are used as substrate. Hysteresis of BChE can be modulated by medium parameters (pH, salts, temperature, and pressure). Though mutant enzymes show different hysteretic behavior, so far attempts to provide a molecular mechanism of BChE hysteresis from mutagenesis studies have been unproductive. However, the substrate dependence of the hysteretic induction times, using wild-type BChE and several mutants, allowed us to build a general, mechanistic model for the hysteresis. In this model, substrate can bind to E, E', or both conformers, and ES and/or E'S can be catalytically active. The exact pathway followed depends on both the nature of the substrate and the structure of the BChE mutant under study. We propose that oscillations develop when substrate exists in different, slowly interconvertible, conformational and/or aggregation forms, of which only the minor form is capable of reacting with BChE. In support of this proposal, NMR studies have provided direct evidence for slow equilibria between monomeric and micellar forms of long-chain, alkyl derivatives of benzoyl-(N-substituted) choline. There is no direct evidence that hysteresis plays a role in BChE function(s). However, the "new view" of protein dynamics proposes that proteins are normally in equilibrium between pre-existing, functional and non-functional conformers; and that binding a ligand to the functional form shifts that equilibrium towards the functional conformation. Therefore, a physiological or toxicological relevance for the hysteresis in BChE cannot be ruled out.

A kinetic model for membrane P-type adenosine triphosphatases is considered, the main application being to the erythrocyte Ca2+-ATPase. It is shown that a simple modification of the known catalytic mechanism of the ATPase by addition of a self-inhibition step and the steady calcium influx leads to damped oscillations in the system discussed. In this way, the model can explain the kinetic experimental results obtained for the purified enzyme in solution as well as for the enzyme incorporated into liposome membranes. The estimated kinetic parameters are close to the experimental ones. Alternative changes in time, demonstrated by the kinetic model for the conformational enzyme states, E(1 )and E(2), confirm the model of two alternatively functioning gates in the ion pumping Ca2+-ATPase.

A kinetic model for the membrane Ca2+-ATPase is considered. The catalytic cycle in the model is extended by enzyme auto-inhibition and by oscillatory calcium influx. It is shown that the conductive enzyme activity can be registered as damped or sustained Ca2+ pulses similar to observed experimentally. It is shown that frequency variations in Ca2+ oscillatory influx induce changes of pulsating enzyme activity. Encoding is observed for the signal frequency into a number of fixed levels of sustained pulses in the enzyme activity. At certain calcium signal frequencies, the calculated Ca2+-ATPase conductivity demonstrates chaotic multi-level pulses, similar to those observed experimentally.

In view of the increasing number of reported concentration oscillations in living cells, methods are needed that can identify the causes of these oscillations. These causes always derive from the influences that concentrations have on reaction rates. The influences reach over many molecular reaction steps and are defined by the detailed molecular topology of the network. So-called 'autoinfluence paths', which quantify the influence of one molecular species upon itself through a particular path through the network, can have positive or negative values. The former bring a tendency towards instability. In this molecular context a new graphical approach is presented that enables the classification of network topologies into oscillophoretic and nonoscillophoretic, i.e. into ones that can and ones that cannot induce concentration oscillations. The network topologies are formulated in terms of a set of uni-molecular and bi-molecular reactions, organized into branched cycles of directed reactions, and presented as graphs. Subgraphs of the network topologies are then classified as negative ones (which can) and positive ones (which cannot) give rise to oscillations. A subgraph is oscillophoretic (negative) when it contains more positive than negative autoinfluence paths. Whether the former generates oscillations depends on the values of the other subgraphs, which again depend on the kinetic parameters. An example shows how this can be established. By following the rules of our new approach, various oscillatory kinetic models can be constructed and analyzed, starting from the classified simplest topologies and then working towards desirable complications. Realistic biochemical examples are analyzed with the new method, illustrating two new main classes of oscillophore topologies.

Steady-state kinetics for the hydrolysis of benzoylcholine (BzCh) and benzoylthiocholine (BzSCh) by wild-type human butyrylcholinesterase (BuChE) and by the peripheral anionic site mutant D70G were compared. kcat/Km for the hydrolysis of BzSCh was 17-fold and 32-fold lower than that for hydrolysis of BzCh by wild-type and D70G, respectively. The rate-limiting step for hydrolysis of BzCh was deacylation, whereas acylation was rate-limiting for hydrolysis of BzSCh. Wild-type enzyme and the D70G mutant were found to reach steady-state velocity slowly with BzCh as the substrate. At pH 6, the approach to steady-state for both enzymes consisted of a mono-exponential acceleration upon which a set of damped oscillations was superimposed. From pH 7 to 8.5, the approach to steady-state consisted of a simple exponential acceleration. The damped oscillations were analyzed by both a numerical approximation and simulation based on a theoretical model. BuChE-catalyzed hydrolysis of the thiocholine analogue of BzCh showed neither lags nor oscillations, under the same conditions. The frequency and amplitude of the damped oscillations decreased as the BzCh concentration increased. The apparent induction time for the exponential portion of the lag was calculated from the envelope of the damped oscillations or from the smooth lag. Wild-type BuChE showed a hyperbolic increase in induction time as the BzCh concentration increased (tau max = 210 s at pH 6.0). However, the induction time for D70G was constant over the whole range of BzCh concentrations (tau max = 60 s at pH 6.0). Thus, the induction time does not conform to a simple hysteretic model in which there is a slow conformational transition of the enzyme from an inactive form E to an active form E'. No pH-dependence of the induction time was found between pH 6.0 and 8.5 in sodium phosphate buffers of various concentrations (from 1 mm to 1 m). However, increasing the pH tended to abolish the oscillations (increase the damping factor). This effect was more pronounced for D70G than for wild-type. Although the lyotropic properties of phosphate change from chaotropic at pH 6.0 to kosmotropic at pH > 8.0, no effect of phosphate concentration on the oscillations was noticed at the different pH values, suggesting that the oscillations are not related to a pH-dependent Hofmeister effect of phosphate ions. Simulation and theoretical analysis of the oscillatory behaviour of the approach to the steady-state for BuChE led us to propose a model for the hysteresis of BuChE with BzCh. In this model, the substrate-free enzyme is present as an equilibrium mixture of two forms, E and E'. Substrate binds to E and E', but only Epsilon'S makes products. It is proposed that oscillations originate from a time-dependent change in the local concentration, solvation and/or conformation of substrate in the bulk solution. 1H-NMR measurements provided evidence for a slow equilibrium between two BzCh conformers. Binding of the conformationally preferred substrate conformer leads to products.

A kinetic model for the bifunctional enzyme, phosphofructo-2-kinase:fructose-2,6-bisphosphatase, is analysed by application of the graph-theoretical method, considering comparable levels for all participants. Certain elementary reactions, distributed on the enzyme surface, are considered to be co-ordinated in a single conformational transition (a model of parallel molecular operations). The method allows us to identify in the kinetic scheme its destabilising sub-scheme as a branched cycle of elementary reactions. Under certain conditions this sub-scheme induces critical phenomena (bistability or oscillations). The computer calculations for the estimated parameter values fit well the experimental observations for this system. The model explains the periodic or bistable counterphase changes of the two opposing activities of this enzyme, observed after glucose perfusion of rat hepatic enzyme samples, and predicts drastic critical changes in kinetic behaviour induced by small external signals. The model also shows the necessity of the phosphoryl intermediate in the mechanism of the bisphosphatase for the critical kind of kinetic behaviour.

The topological structure of the simplest critical fragments in biochemical systems has been characterized. The conditions are considered where the critical fragments induce oscillations of the concentrations of the system participants. To illustrate, three biochemical systems (transport of ions through a membrane, protein phosphorylation, and two-substrate reaction) are discussed. The kinetic schemes of these systems contain one of the discovered critical fragments. Relaxation oscillations of the concentrations of the system participants were demonstrated using the numerical integration method.

The dependence of pyruvate dehydrogenase complex (PDC) activity on [Ca2+] was determined in Ehrlich ascites carcinoma cells at different pyruvate concentrations. The resulting family of curves had the following characteristics: a) bell-shaped appearance of all curves with maximum activity at 600 nM Ca2+; b) unchanged position of maxima with changes in pyruvate concentration; c) nonmonotonous changes in PDC activity with increasing pyruvate concentration at fixed [Ca2+]. Feasible mechanisms involving Ca2+-dependent phosphatase and kinase which are consistent with the experimental findings are discussed. To determine the steps in the chain of PDC reactions which determine the observed phenomena, a mathematical model is suggested which is based on the known data on the structural--functional relationships between the complex components--pyruvate dehydrogenase (E1), dihydrolipoyl acetyl transferase (E2), dihydrolipoyl dehydrogenase (E3), protein X, kinase, and phosphatase. To adequately describe the non-trivial dependence of PDC activity on [Ca2+] at different pyruvate concentrations, it was also necessary to consider the interdependence of some steps in the general chain of PDC reactions. Phenomenon (a) is shown to be due only to the involvement of protein X in the PDC reactions, phenomenon (b) to be due to changes in the activity of kinase, and phenomenon (c) to be due to dependence of acetylation and transacetylation rates on pyruvate concentration.

Recent experimental findings on the structural--functional features of pyruvate dehydrogenase phosphatase (PDP) isolated from various sources are compared. Two alternative mechanisms (a and b) of dephosphorylation of the E1 component in the pyruvate dehydrogenase complex (PDC) are discussed: a) the reaction occurs as a result of stochastic collisions of PDP and PDC, and the generation of an enzyme--substrate complex (PDP--E1--PDC) and dephosphorylation of the E1 component occur independently at different PDP binding sites on the PDC core; b) the dephosphorylation is performed simultaneously by a certain number of PDP molecules symmetrically bound on the PDC core. The second mechanism is suggested by the self-assembly theory of multicomponent enzyme systems and can be proved by kinetic experiments. Based on self-assembly principles and data on feasible binding sites of peripheral components of the PDC, the stoichiometry and mutual location of PDP, pyruvate dehydrogenase kinase, and the E1 component on the core of mammalian PDC are postulated to provide optimal functioning of the PDC. Structural mechanisms of stimulation of PDP activity by Ca2+ and polyamines are also discussed.

The structure-function organization of mammalian pyruvate dehydrogenase complex (PDC) is considered. The linear size of the complex components in relation to the size of their fragments and mean distances between potentially active sites are estimated. A kinetic model for the complex with core subunits divided into conformational classes characterized by their different activities is discussed. The kinetic features of this model are compared with the features of the model for the core subunits not divided into classes. Curves are described for the activity of PDC versus time for various activities of the kinase and the phosphatase as dependent on their binding to more or less active core subunits. The results show that spatially distributed multiple active sites in the multi-enzyme complex can drastically change their full activity in time without any marked changes in the rate constants of the elementary reaction steps due only to spatially changed reaction pathways. A new type of regulation discussed for PDC reveals unusual regulatory abilities for complexes of this family.

The experimentally observed phenomenon of non-equimolarity for enzyme components, assembled into multienzyme complexes of the 2-oxo acid dehydrogenases family, is structurally interpreted to predict the only possible stable symmetrical distribution of peripheral components on the complex core. To obey the equivalent neighboring, that is necessary for unique self-assembled structures, we should deduce discrete conformational states for core subunits, those with different affinity for peripheral components. Two kinetically different types of substrate-intermediate pathways through the lipoyl network of the mammalian pyruvate dehydrogenase complex follow from this structural theory. The theory predicts unusual kinetic behavior for the multienzyme complex.

A kinetic model for the pyruvate dehydrogenase complex is analyzed. The model takes into account intermediate channeling through the lipoyl network attached to the complex core, as well as inter-related regulatory effects of protein X acetylation and enzyme phosphorylation. The model predicts undamped oscillations of enzyme activity.

A novel regulation type, which may be observed as an unusual kinetic 'cooperativity', is predicted for the alpha-keto acid dehydrogenase complexes. The inter-relationship of this regulation with the well-known regulatory effect of enzyme phosphorylation is discussed.

A graph-theoretic approach is shown to be applicable within the framework of the metabolic control analysis. Kinetic differential equations linearized near a steady state are presented as kinetic graphs (schemes), their structure being correlated with kinetic properties of corresponding metabolic networks. The global properties may be expressed in terms of the local properties for steady states of metabolic systems. Instability, bistability, and concentrational oscillations are shown to be induced by specific graph fragments. The approach is illustrated by an example of systems showing the oscillatory kinetic behaviour.

Stability of multienzyme systems with feedback regulation has been analyzed on the basis of the Lienard-Chipart criteria. The rules governing the topological graph construction for multienzyme systems have been developed. A theorem about correspondence of the graph constructed and coefficients of the characteristic polynomial of linearized kinetic equations is proved. The graph-theoretical stability analysis proposed is illustrated by a number of examples of multienzyme systems with feedback regulation.

A graphical method to reveal the so-called 'critical fragments' in schemes of biochemical systems is considered. These fragments produce multiple steady states or self-oscillations in systems. As an example, the bifunctional enzyme, 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase, regulated by glucagon through enzyme phosphorylation, is discussed. It is shown that this enzyme may act as a metabolic switching mechanism in discontinuous or oscillatory regimes, depending on the specific structure of its kinetic scheme. The boundaries of concentrational and parameter domains for these critical phenomena are also predicted.

The development of the graph-theoretic method is proposed particularly for the analysis of closed cycles of elementary stages in enzyme reaction schemes. Some simplifications of the graph structure may be based on the application of Kirchhoff's laws to enzyme reaction graphs in the steady-state. The importance of the cyclic processes for enzyme regulations and a principle non-equilibrium of this phenomenon are emphasized. As an example of the regulatory role of cycles "the liberation" from substrate inhibition by substrate analogues is considered. The modification of the graph-theoretic method in the pre-steady-state kinetics for arbitrary initial conditions (for pre-mixing procedures) is also discussed. The necessary and sufficient conditions for damped oscillations in the pre-steady state are formulated which are the equality conditions for some of the rate constants along the cycle (both for reversible and irreversible stages).