Neurotoxic and convulsant effects induced by jack bean ureases on the mammalian nervous system.

Ureases are microbial virulence factors either because of the enzymatic release of ammonia or due to many other non-enzymatic effects. Here we studied two neurotoxic urease isoforms, Canatoxin (CNTX) and Jack Bean Urease (JBU), produced by the plant Canavalia ensiformis, whose mechanisms of action remain elusive. The neurotoxins provoke convulsions in rodents (LD50 ∼2 mg/kg) and stimulate exocytosis in cell models, affecting intracellular calcium levels. Here, electrophysiological and brain imaging techniques were applied to elucidate their mode of action. While systemic administration of the toxins causes tonic-clonic seizures in rodents, JBU injected into rat hippocampus induced spike-wave discharges similar to absence-like seizures. JBU reduced the amplitude of compound action potential from mouse sciatic nerve in a tetrodotoxin-insensitive manner. Hippocampal slices from CNTX-injected animals or slices treated in vitro with JBU failed to induce long term potentiation upon tetanic stimulation. Rat cortical synaptosomes treated with JBU released L-glutamate. JBU increased the intracellular calcium levels and spontaneous firing rate in rat hippocampus neurons. MicroPET scans of CNTX-injected rats revealed increased [18]Fluoro-deoxyglucose uptake in epileptogenesis-related areas like hippocampus and thalamus. Curiously, CNTX did not affect voltage-gated sodium, calcium or potassium channels currents, neither did it interfere on cholinergic receptors, suggesting an indirect mode of action that could be related to the ureases' membrane-disturbing properties. Understanding the neurotoxic mode of action of C. ensiformis ureases could help to unveil the so far underappreciated relevance of these toxins in diseases caused by urease-producing microorganisms, in which the human central nervous system is affected.

Soluble Epoxide Hydrolase and Brain Cholesterol Metabolism.

The bifunctional enzyme soluble epoxide hydrolase (sEH) is found in all regions of the brain. It has two different catalytic activities, each assigned to one of its terminal domains: the C-terminal domain presents hydrolase activity, whereas the N-terminal domain exhibits phosphatase activity. The enzyme's C-terminal domain has been linked to cardiovascular protective and anti-inflammatory effects. Cholesterol-related disorders have been associated with sEH, which plays an important role in the metabolism of cholesterol precursors. The role of sEH's phosphatase activity has been so far poorly investigated in the context of the central nervous system physiology. Given that brain cholesterol disturbances play a role in the onset of Alzheimer's disease (AD) as well as of other neurodegenerative diseases, understanding the functions of this enzyme could provide pivotal information on the pathophysiology of these conditions. Moreover, the sEH phosphatase domain could represent an underexplored target for drug design and therapeutic strategies to improve symptoms related to neurodegenerative diseases. This review discusses the function of sEH in mammals and its protein structure and catalytic activities. Particular attention was given to the distribution and expression of sEH in the human brain, deepening into the enzyme's phosphatase activity and its participation in brain cholesterol synthesis. Finally, this review focused on the metabolism of cholesterol and its association with AD.

Ureases: Historical aspects, catalytic, and non-catalytic properties - A review.

Urease (urea amidohydrolase, EC 3.5.1.5) is a nickel-containing enzyme produced by plants, fungi, and bacteria that catalyzes the hydrolysis of urea into ammonia and carbamate. Urease is of historical importance in Biochemistry as it was the first enzyme ever to be crystallized (1926). Finding nickel in urease's active site (1975) was the first indication of a biological role for this metal. In this review, historical and structural features, kinetics aspects, activation of the metallocenter and inhibitors of the urea hydrolyzing activity of ureases are discussed. The review also deals with the non-enzymatic biological properties, whose discovery 40 years ago started a new chapter in the study of ureases. Well recognized as virulence factors due to the production of ammonia and alkalinization in diseases by urease-positive microorganisms, ureases have pro-inflammatory, endocytosis-inducing and neurotoxic activities that do not require ureolysis. Particularly relevant in plants, ureases exert insecticidal and fungitoxic effects. Data on the jack bean urease and on jaburetox, a recombinant urease-derived peptide, have indicated that interactions with cell membrane lipids may be the basis of the non-enzymatic biological properties of ureases. Altogether, with this review we wanted to invite the readers to take a second look at ureases, very versatile proteins that happen also to catalyze the breakdown of urea into ammonia and carbamate.

Peptide YY (3-36) modulates intracellular calcium through activation of the phosphatidylinositol pathway in hippocampal neurons.

Peptide YY (PYY) belongs to the neuropeptide Y (NPY) family, which also includes the pancreatic polypeptide (PP) and NPY. PYY is secreted by the intestinal L cells, being present in the blood stream in two active forms capable of crossing the blood brain barrier, PYY (1-36) and its cleavage product, PYY (3-36). PYY is a selective agonist for the Y2 receptor (Y2R) and these receptors are abundant in the hippocampus. Here we investigated the mechanisms by which PYY (3-36) regulates intracellular Ca2+ concentrations ([Ca2+]i) in hippocampal neurons by employing a calcium imaging technique in hippocampal cultures. Alterations in [Ca2+]i were detected by changes in the Fluo-4 AM reagent emission. PYY (3-36) significantly increased [Ca2+] from the concentration of 10-11M as compared to the controls (infusion of HEPES-buffered solution (HBS) solution alone). The PYY (3-36)-increase in [Ca2+]i remained unchanged even in Ca2+-free extracellular solutions. Sarcoplasmic/endoplasmic reticulum Ca2+-ATPase pump (SERCA pump) inhibition partially prevent the PYY (3-36)-increase of [Ca2+]i and inositol 1,4,5-triphosphate receptor (IP3R) inhibition also decreased the PYY (3-36)-increase of [Ca2+]i. Taken together, our data strongly suggest that PYY (3-36) mobilizes calcium from the neuronal endoplasmic reticulum (ER) stores towards the cytoplasm. Next, we showed that PYY (3-36) inhibited high K+-induced increases of [Ca2+]i, suggesting that PYY (3-36) could also act by activating G-protein coupled inwardly rectifying potassium K+ channels. Finally, the co-infusion of the Y2 receptor (Y2R) antagonist BIIE0246 with PYY (3-36) abolished the [Ca2+]i increase induced by the peptide, suggesting that PYY (3-36)-induced [Ca2+]i increase in hippocampal neurons occurs via Y2Rs.

Methylprednisolone as a memory enhancer in rats: Effects on aversive memory, long-term potentiation and calcium influx.

It is well recognized that stress or glucocorticoids hormones treatment can modulate memory performance in both directions, either impairing or enhancing it. Despite the high number of studies aiming at explaining the effects of glucocorticoids on memory, this has not yet been completely elucidated. Here, we demonstrate that a low daily dose of methylprednisolone (MP, 5mg/kg, i.p.) administered for 10-days favors aversive memory persistence in adult rats, without any effect on the exploring behavior, locomotor activity, anxiety levels and pain perception. Enhanced performance on the inhibitory avoidance task was correlated with long-term potentiation (LTP), a phenomenon that was strengthen in hippocampal slices of rats injected with MP (5mg/kg) during 10days. Additionally, in vitro incubation with MP (30-300µM) concentration-dependently increased intracellular [Ca2+]i in cultured hippocampal neurons depolarized by KCl (35mM). In conclusion, a low daily dose of MP for 10days may promote aversive memory persistence in rats.

Bothriurus bonariensis scorpion venom activates voltage-dependent sodium channels in insect and mammalian nervous systems.

Animal venoms have been widely recognized as a major source of biologically active molecules. Bothriurus bonariensis, popularly known as black scorpion, is the arthropod responsible for the highest number of accidents involving scorpion sting in Southern Brazil. Here we reported the first attempt to investigate the neurobiology of B. bonariensis venom (BBV) in the insect and mammalian nervous system. BBV (32 µg/g) induced a slow neuromuscular blockade in the in vivo cockroach nerve-muscle preparations (70 ± 4%, n = 6, p < 0.001), provoking repetitive twitches and significantly decreasing the frequency of spontaneous leg action potentials (SNCAPs) from 82 ± 3 min(-1) to 36 ± 1.3 min(-1) (n = 6, p < 0.05), without affecting the amplitude. When tested in primary cultures of rat hippocampal cells, BBV induced a massive increase of Ca(2+) influx (250 ± 1% peak increase, n = 3, p < 0.0001). The disturbance of calcium homeostasis induced by BBV on the mammalian central nervous system was not accompanied by cellular death and was prevented by the co-treatment of the hippocampal cells with tetrodotoxin, a selective sodium channel blocker. The results suggest that the biological activity of BBV is mostly related to a modulation of sodium channels function. Our biological activity survey suggests that BBV may have a promising insecticidal and therapeutic potential.