Magnesium and Morphine in the Treatment of Chronic Neuropathic Pain-A Biomedical Mechanism of Action.

The effectiveness of opioids in the treatment of neuropathic pain is limited. It was demonstrated that magnesium ions (Mg2+), physiological antagonists of N-methyl-D-aspartate receptor (NMDAR), increase opioid analgesia in chronic pain. Our study aimed to determine the molecular mechanism of this action. Early data indicate the cross-regulation of µ opioid receptor (MOR) and NMDAR in pain control. Morphine acting on MOR stimulates protein kinase C (PKC), while induction of NMDAR recruits protein kinase A (PKA), leading to a disruption of the MOR-NMDAR complex and promoting functional changes in receptors. The mechanical Randall-Selitto test was used to assess the effect of chronic Mg2+ and morphine cotreatment on streptozotocin-induced hyperalgesia in Wistar rats. The level of phosphorylated NMDAR NR1 subunit (pNR1) and phosphorylated MOR (pMOR) in the periaqueductal gray matter was determined with the Western blot method. The activity of PKA and PKC was examined by standard enzyme immunoassays. The experiments showed a reduction in hyperalgesia after coadministration of morphine (5 mg/kg intraperitoneally) and Mg2+ (40 mg/kg intraperitoneally). Mg2+ administered alone significantly decreased the level of pNR1, pMOR, and activity of both tested kinases. The results suggest that blocking NMDAR signaling by Mg2+ restores the MOR-NMDAR complex and thus enables morphine analgesia in neuropathic rats.

GABAB receptor intracellular signaling: novel pathways for depressive disorder treatment?

Affecting over 320 million people around the world, depression has become a formidable challenge for modern medicine. In addition, an increasing number of studies cast doubt on the monoamine theory of depressive disorder and, worryingly, antidepressant medications only significantly benefit patients with severe depression. Thus, it is not surprising that researchers have shown an increased interest in new theories attempting to explain the pathogenesis of this disease. One example is the excitatory/inhibitory transmission imbalance theory. These abnormalities involve glutamate and γ-aminobutyric acid (GABA) signaling. Studies on GABAB receptors and their antagonists are particularly promising for the treatment of depressive disorders. In this paper, intracellular pathways controlled by GABAB receptors and their links to depression are described, including the impact of ketamine on GABAergic synaptic transmission.

Ionic Mechanism Underlying Rebound Depolarization in Medial Prefrontal Cortex Pyramidal Neurons.

Rebound depolarization (RD) occurs after membrane hyperpolarization and converts an arriving inhibitory signal into cell excitation. The purpose of our study was to clarify the ionic mechanism of RD in synaptically isolated layer V medial prefrontal cortex (mPFC) pyramidal neurons in slices obtained from 58- to 62-day-old male rats. The RD was evoked after a step hyperpolarization below -80 mV, longer than 150 ms in 192 of 211 (91%) tested neurons. The amplitude of RD was 30.6 ± 1.2 mV above the resting membrane potential (-67.9 ± 0.95 mV), and it lasted a few 100 ms (n = 192). RD could be observed only after preventing BK channel activation, which was attained either by using paxilline, by removal of Ca++ from the extra- or intracellular solution, by blockade of Ca++ channels or during protein kinase C (PKC) activation. RD was resistant to tetrodotoxin (TTX) and was abolished after the removal of Na+ from the extracellular solution or application of an anti-Nav1.9 antibody to the cell interior. We conclude that two membrane currents are concomitantly activated after the step hyperpolarization in the tested neurons: a. a low-threshold, TTX-resistant, Na+ current that evokes RD; and b. an outward K+ current through BK channels that opposes Na+-dependent depolarization. The obtained results also suggest that a. low-level Ca++ in the external medium attained upon intense neuronal activity may facilitate the formation of RD and seizures; and b. RD can be evoked during the activation of PKC, which is an effector of a number of transduction pathways.

Noradrenaline Modulates the Membrane Potential and Holding Current of Medial Prefrontal Cortex Pyramidal Neurons via ß1-Adrenergic Receptors and HCN Channels.

The medial prefrontal cortex (mPFC) receives dense noradrenergic projections from the locus coeruleus. Adrenergic innervation of mPFC pyramidal neurons plays an essential role in both physiology (control of memory formation, attention, working memory, and cognitive behavior) and pathophysiology (attention deficit hyperactivity disorder, posttraumatic stress disorder, cognitive deterioration after traumatic brain injury, behavioral changes related to addiction, Alzheimer's disease and depression). The aim of this study was to elucidate the mechanism responsible for adrenergic receptor-mediated control of the resting membrane potential in layer V mPFC pyramidal neurons. The membrane potential or holding current of synaptically isolated layer V mPFC pyramidal neurons was recorded in perforated-patch and classical whole-cell configurations in slices from young rats. Application of noradrenaline (NA), a neurotransmitter with affinity for all types of adrenergic receptors, evoked depolarization or inward current in the tested neurons irrespective of whether the recordings were performed in the perforated-patch or classical whole-cell configuration. The effect of noradrenaline depended on ß1- and not α1- or α2-adrenergic receptor stimulation. Activation of ß1-adrenergic receptors led to an increase in inward Na+ current through hyperpolarization-activated cyclic nucleotide-gated (HCN) channels, which carry a mixed Na+/K+ current. The protein kinase A- and C-, glycogen synthase kinase-3ß- and tyrosine kinase-linked signaling pathways were not involved in the signal transduction between ß1-adrenergic receptors and HCN channels. The transduction system operated in a membrane-delimited fashion and involved the ßγ subunit of G-protein. Thus, noradrenaline controls the resting membrane potential and holding current in mPFC pyramidal neurons through ß1-adrenergic receptors, which in turn activate HCN channels via a signaling pathway involving the ßγ subunit.

Age-dependent expression of Nav1.9 channels in medial prefrontal cortex pyramidal neurons in rats.

Developmental changes that occur in the prefrontal cortex during adolescence alter behavior. These behavioral alterations likely stem from changes in prefrontal cortex neuronal activity, which may depend on the properties and expression of ion channels. Nav1.9 sodium channels conduct a Na+ current that is TTX resistant with a low threshold and noninactivating over time. The purpose of this study was to assess the presence of Nav1.9 channels in medial prefrontal cortex (mPFC) layer II and V pyramidal neurons in young (20-day old), late adolescent (60-day old), and adult (6- to 7-month old) rats. First, we demonstrated that layer II and V mPFC pyramidal neurons in slices obtained from young rats exhibited a TTX-resistant, low-threshold, noninactivating, and voltage-dependent Na+ current. The mRNA expression of the SCN11a gene (which encodes the Nav1.9 channel) in mPFC tissue was significantly higher in young rats than in late adolescent and adult rats. Nav1.9 protein was immunofluorescently labeled in mPFC cells in slices and analyzed via confocal microscopy. Nav1.9 immunolabeling was present in layer II and V mPFC pyramidal neurons and was more prominent in the neurons of young rats than in the neurons of late adolescent and adult rats. We conclude that Nav1.9 channels are expressed in layer II and V mPFC pyramidal neurons and that Nav1.9 protein expression in the mPFC pyramidal neurons of late adolescent and adult rats is lower than that in the neurons of young rats. © 2017 Wiley Periodicals, Inc. Develop Neurobiol 77: 1371-1384, 2017.

Immunological factors and liver fibrosis in pediatric liver transplant recipients.

BACKGROUND: The aim of our study was to retrospectively assess any correlation between graft fibrosis and selected immunological factors in pediatric liver transplant recipients. MATERIAL AND METHODS: The study was performed on 33 patients after living related donor transplantation, divided into 2 groups depending on history of acute rejection episodes after transplantation. We assessed liver biopsies for presence of fibrosis, signs of antibody-mediated rejection, inflammatory infiltrations, and changes in bile ducts. We correlated these findings with assessment of anti-HLA antibodies. RESULTS: Among 14 patients with ACR, a history fibrosis was found in 8 patients (57%). In 19 patients without a history of ACR, fibrosis was found in 9 patients (47%). Anti-HLA antibodies were found in 47% of patients with fibrosis and in only 18.75% of patients without fibrosis. Among 3 patients with signs of antibody-mediated rejection, all had fibrosis in the graft 2 years after transplantation. We did not find any patient with chronic rejection or ductopenia. CONCLUSIONS: We suggest that there is a correlation between ACR and development of graft fibrosis present in liver grafts from recipients with normal liver biochemistry. Anti-HLA antibodies class II seems to be most important in development of fibrosis.

Antibody-mediated rejection in pediatric liver transplant recipients.

BACKGROUND: Antibody-mediated rejection (AMR), associated with the presence of C4d deposits, is well-defined in kidney transplantation but much less documented in liver transplantation (LTx). The aim of our study was to retrospectively analyze a group of pediatric liver transplant recipients who experienced episodes of acute rejection in the past, for the signs of AMR and its impact on liver histology. MATERIAL/METHODS: Our study population consisted of 18 patients after living related donor liver transplantation with a history of acute cellular rejection (1-5/patient). In all of them, actual liver function was good at almost 2-year median follow-up after transplantation. We reassessed all liver biopsies taken from these children between 5 days to 5.7 years after transplantation for signs of acute cellular rejection and antibody-mediated rejection. In all patients, anti-HLA antibodies were also assessed at least 2 years after transplantation (2.18-12.27 years, median 6.795 years). RESULTS: There were 27 episodes of acute rejection proved by liver biopsy. Signs of AMR were found in 6 of 18 patients (33.3%). In 5 of these patients, donor-specific (DSA) and non-specific anti-HLA antibodies were also identified. In the group of 12 patients with acute rejection without histochemical signs of AMR, anti-HLA antibodies were found in sera of only 5 of 12 patients after transplantation. CONCLUSIONS: Our study shows some correlation between C4d-positive reaction in liver biopsies with acute cellular rejection and presence of anti-HLA antibodies, particularly against HLA class II. We did not find any difference in the late graft function, which could be correlated with the presence of AMR. Further studies on larger groups of patients are necessary.

Significant changes in the composition of the precursor B-cell compartment in children less than 2 years old.

BACKGROUND: Defects in early B lymphocyte maturation in bone marrow (BM) compose a characteristic feature of many primary immune deficiencies associated with agammaglobulinemia. To date, only limited data on the composition of the precursor B-cell compartment in BM is available. The aim of this study was to define normal age-related ranges of total B-cell content and distribution of precursor B-cell stages in BM for the future use in clinical diagnostics. METHODS: Four color flow cytometry was used to analyze the composition of the B-cell compartment in specimens from 59 hematologically healthy children, aged 14 days to 16 years, assigned to six age groups: neonates less than 1 month old, infants >1-12 months old, children >1-2 years old, >2-5 years old, >5-10 years old, and older than 10 years. RESULTS: Analysis of the composition of the B-cell compartment revealed significant age-related variation in the distribution of individual B-cell maturation stages, most seriously affecting children during first 2 years of life, with the shift from domination of the earliest stages, to gradually increasing content of mature B-cells. Significantly higher proportions of pro-B lymphocytes were observed in neonates than in any other age group. CONCLUSION: Physiological age-related variation in the precursor B-cell compartment composition affects most seriously very young children below the age of 2 years. Proper interpretation of immunophenotyping results performed in cases of suspected early B-cell differentiation defect requires application of adequate reference data.