Transmission of Donor-Derived Trypanosoma cruzi and Subsequent Development of Chagas Disease in a Lung Transplant Recipient.

Donor infection status should be considered when accepting an organ for transplant. Here we present a case of Chagas disease developing after a lung transplant where the donor was known to be Trypanosoma cruzi antibody positive. The recipient developed acute Trypanosoma cruzi infection with reactivation after treatment. Chagas disease-positive donors are likely to be encountered in the United States; donor targeted screening is needed to guide decisions regarding organ transplant and posttransplant monitoring.

Neuron-microglia communication in the CNS of the freshwater snail Planorbarius corneus.

The aim of the present study was to identify molecules that may be involved in neuron-microglia communication in the CNS of freshwater snail Planorbarius corneus. Messenger molecules are exchanged in normal and pathological conditions and we tried to identify some of them by immunocytochemistry on whole ganglia and cell cultures. In particular, we examined neurons and microglia for the expression of some cytokines, IL-1alpha, IL-1beta, IL-6 and TNF-alpha and the neurotransmitter glutamate. These substances may be released by suffering or injured neurons and communicate to microglia the damaging event. Even microglia, on own turn, once activated, express and released the same or other substances in order to reestablish the system homeostasis, depending on modalities and times of activation. We discuss the possibility that hyperactivated microglia can shift from neuroprotective to neurodegenerative. Moreover, we examined in neuron-microglia co-coltures the direct interaction effects in terms of neuronal survival and improved neurite regeneration.

Microglia proliferation as a response to activation in the freshwater snail Planorbarius corneus: a BrdU incorporation study.

Invertebrate microglia constitute a class of cells resident in the ganglionic nervous system which are activated after tissue injury or by the presence of pathogens. The microglia activation response includes graduated morpho-functional and biochemical changes and cell proliferation. In this study we verified in the freshwater snail Planorbarius corneus that an activation caused by a traumatic event may induce microglia division. Cell proliferation was assessed immunocytochemically using BrdU incorporation technique and documented on both ganglionic sections and microglia cultured cells at different experimental conditions and times after activation. In addition, we studied the possibility of increasing microglia proliferation by adding to the cultured medium the Macrophage-Colony Stimulating Factor (M-CSF) that has been shown to stimulate specifically this process in vertebrates.

Endogenous morphine modulates acute thermonociception in mice.

The endogenous synthesis of morphine has been clearly demonstrated throughout the phylogenesis of the nervous system of mammals and lower animals. Endogenous morphine, serving as either a neurotransmitter or neurohormone, has been demonstrated in the nervous system of both vertebrates and invertebrates. As one of the effects of exogenous morphine is the modulation of pain perception, we investigated the effects that the depletion of endogenous morphine had on nociceptive transmission. The immunoneutralization of endogenous morphine from brain extracellular spaces was obtained through the intracerebroventricular administration of affinity purified anti-morphine IgG to mice, which then underwent the hot plate test. Endogenous morphine immunoneutralization decreased thermal response latency and attenuated the anti-nociceptive effect of the mu selective agonist DAMGO in hot plate test suggesting that endogenous morphine is involved in pain modulation.

Effects of respiratory muscle training versus placebo on endurance exercise performance.

We evaluated the effects of a 5 week (25 sessions); (30-35 min/day, 5 days/week), respiratory muscle training (RMT) program in nine competitive male cyclists. The experimental design included inspiratory resistance strength training (3-5 min/session) and hyperpnea endurance training (30 min/session), a placebo group which used a sham hypoxic trainer (n=8), and three exercise performance tests, including a highly reproducible 8 km time trial test. RMT intensity, measured once a week in terms of accumulated inspiratory pressure and the level of sustainable hyperpnea increased significantly after 5 weeks (+64% and +19%, respectively). The RMT group showed a significant 8% increase in maximal inspiratory pressure (P<0.05) while the placebo group showed only a 3.7% increase (P>0.10). RMT and placebo groups both showed significant increases in the fixed work-rate endurance test performance time (+26% and +16%, respectively) and in the peak work-rate achieved during the incremental maximal oxygen consumption (V(O2)max) test (+9 and +6%). The 8 km time trial performance increased 1.8+/-1.2% (or 15+/-10 sec; P<0.01) in the RMT group with 8 of 9 subjects increasing; the placebo group showed a variable non-significant change in 5 of 8 subjects (-0.3+/-2.7%, P=0.07). The changes observed in these three performance tests were not, however, significantly different between the RMT and placebo groups. Heart rate, ventilation, or venous blood lactate, at equal work-rates during the incremental exercise test or at equal times during the fixed work-rate endurance test were not changed significantly across these exercise trials in either group. We propose that the effect of RMT on exercise performance in highly trained cyclists does not exceed that in a placebo group. Significant placebo and test familiarization effects must be accounted for in experimental designs utilizing performance tests which are critically dependent on volitional effort.

Effects of exhaustive endurance exercise on pulmonary gas exchange and airway function in women.

Seventeen fit women ran to exhaustion (14 +/- 4 min) at a constant speed and grade, reaching 95 +/- 3% of maximal O(2) consumption. Pre- and postexercise lung function, including airway resistance [total respiratory resistance (Rrs)] across a range of oscillation frequencies, was measured, and, on a separate day, airway reactivity was assessed via methacholine challenge. Arterial O(2) saturation decreased from 97.6 +/- 0.5% at rest to 95.1 +/- 1.9% at 1 min and to 92.5 +/- 2.6% at exhaustion. Alveolar-arterial O(2) difference (A-aDO(2)) widened to 27 +/- 7 Torr after 1 min and was maintained at this level until exhaustion. Arterial PO(2) (Pa(O(2))) fell to 80 +/- 8 Torr at 1 min and then increased to 86 +/- 9 Torr at exhaustion. This increase in Pa(O(2)) over the exercise duration occurred due to a hyperventilation-induced increase in alveolar PO(2) in the presence of a constant A-aDO(2). Arterial O(2) saturation fell with time because of increasing temperature (+2.6 +/- 0.5 degrees C) and progressive metabolic acidosis (arterial pH: 7.39 +/- 0.04 at 1 min to 7.26 +/- 0.07 at exhaustion). Plasma histamine increased throughout exercise but was inversely correlated with the fall in Pa(O(2)) at end exercise. Neither pre- nor postexercise Rrs, frequency dependence of Rrs, nor diffusing capacity for CO correlated with the exercise A-aDO(2) or Pa(O(2)). Although several subjects had a positive or borderline hyperresponsiveness to methacholine, this reactivity did not correlate with exercise-induced changes in Rrs or exercise-induced arterial hypoxemia. In conclusion, regardless of the degree of exercise-induced arterial hypoxemia at the onset of high-intensity exercise, prolonging exercise to exhaustion had no further deleterious effects on A-aDO(2), and the degree of gas exchange impairment was not related to individual differences in small or large airway function or reactivity.

Endogenous morphine.

It is now well accepted that endogenous morphine is present in animals, both in invertebrates and vertebrates. It is a key signaling molecule that plays an important role in downregulating physiological responses, such as those in the immune system, including immune elements in the CNS. It has been demonstrated that a specific mu-opiate-receptor subtype, mu3, mediates these downregulatory effects through release of NO. This article examines morphine as an endogenous signaling molecule, in terms of its role in neural and immune regulation.

Ascaris suum, an intestinal parasite, produces morphine.

The parasitic worm Ascaris suum contains the opiate alkaloid morphine as determined by HPLC coupled to electrochemical detection and by gas chromatography/mass spectrometry. The level of this material is 1168 +/- 278 ng/g worm wet weight. Furthermore, Ascaris maintained for 5 days contained a significant amount of morphine, as did their medium, demonstrating their ability to synthesize the opiate alkaloid. To determine whether the morphine was active, we exposed human monocytes to the material, and they immediately released nitric oxide in a naloxone-reversible manner. The anatomic distribution of morphine immunoreactivity reveals that the material is in the subcuticle layers and in the animals' nerve chords. Furthermore, as determined by RT-PCR, Ascaris does not express the transcript of the neuronal mu receptor. Failure to demonstrate the expression of this opioid receptor, as well as the morphine-like tissue localization in Ascaris, suggests that the endogenous morphine is intended for secretion into the microenvironment.

Endogenous morphine levels increase in molluscan neural and immune tissues after physical trauma.

The aim of this study was to demonstrate by biochemical and immunocytochemical methods the presence of endogenous morphine in nervous and immune tissues of the freshwater snail, Planorbarius corneus. High performance liquid chromatography (HPLC) coupled to electrochemical detection performed on tissues from control snails, revealed that the CNS contains 6.20+/-2.0 pmol/g of the alkaloid, the foot tissue contains a much lower level, 0.30+/-0.03 pmol/g, whilst morphine is not detected in the hemolymph and hepatopancreas. In specimens that were traumatized, we detected a significant rise of the CNS morphine level 24 h later (43.7+/-5.2 pmol/g) and an initial decrease after 48 h (19.3+/-4.6 pmol/g). At the same times, we found the appearance of the opiate in the hemolymph (0.38+/-0.04 pmol/ml and 0.12+/-0.03 pmol/ml) but not in the hepatopancreas. Using indirect immunocytochemistry, a morphine-like molecule was localized to a number of neurons and a type of glial cell in the CNS, to some immunocytes in the hemolymph and to amoebocytes in the foot, as well as to fibers in the aorta wall. Simultaneously to the rise of morphine biochemical level following trauma, morphine-like immunoreactivity (MIR) increased in both intensity and the number of structures responding positively, i.e., neurons and fiber terminals. In another mollusc, the mussel Mytilus galloprovincialis, the same pattern of enhanced MIR was found after trauma. Taken together, the data suggest the presence of a morphinergic signaling in invertebrate neural and immune processes resembling those of classical messenger systems and an involvement in trauma response.

Opiate signaling regulates microglia activities in the invertebrate nervous system.

1. Evidence supporting the presence in the invertebrate nervous system of a class of glial cells resembling vertebrate microglia was obtained in the freshwater snail Planorbarius corneus. These cells are easily identified by their immunopositivity to anti-pro-opiomelanocortin (POMC)-derived peptide antibodies. 2. Invertebrate microglia, as in vertebrates, exhibit macrophage-like activity in vivo and in cell cultures. These cells respond to the trauma of ganglionic excision and their organotypic culture by leaving their location around neurons and moving to the lesion site from which they migrate in the culture dish. 3. In vitro, these microglia undergo conformational changes and show phagocytic properties in the presence of bacteria or lipopolysaccharide. The activated cells also express tumor necrosis factor-alpha-like material and an increase in nitric oxide synthase, as shown by immunocytochemistry. 4. The inhibitory effect of morphine on the mobility and phagocytic activity of invertebrate microglia provide additional functional evidence for a possible role of opiate-like compounds in downregulating immunoregulatory processes, as also observed in the circulating immunocytes.

Effect of serotonin and neuropeptides on adenylate cyclase of the central nervous system and peripheral organs of the freshwater snail Planorbarius corneus.

The effect of serotonin, FMRFamide and the small cardioactive peptide B (SCPB) on adenylate cyclase activity of the central nervous system and some peripheral organs of the freshwater snail Planorbarius corneus was investigated. The amine and the cardioactive peptide stimulated the enzyme, although with different potencies, in all tissues studied and, when tested in combination, an additive activation was obtained. FMRFamide induced differential effects in the various targets: marked stimulation of adenylate cyclase, additive to that provoked by serotonin or SCPB, in salivary glands; inhibition of the enzyme, both alone and in combination with the other neuromediators, in the nervous tissue; whereas no influence was found in adenylate cyclase activity in the buccal mass. In the last of these tissues, the peptide might act through an intracellular second messenger other than cyclic AMP. The responsiveness of adenylate cyclase to these neuromediators in all the central ganglia suggested that they can exert an important role as neurotransmitters and/or neuromodulators in the central nervous system of the snail. Moreover, in the light of the differential sensitivity of adenylate cyclase in the salivary glands and buccal mass, we suggest that serotonin, FMRFamide and SCPB modulate the feeding behaviour of P. corneus in a complex way.

MDL 12330A inhibits the non-neuronal adenylyl cyclase from the freshwater snail Planorbarius corneus, but the neuronal enzyme is activated by this compound.

N-(Cis-2-phenyl-cyclopentyl)azacyclotridecan-2-imine-hydrochloride (MDL 12330A), considered an inhibitor of adenylyl cyclase, has been tested on the enzyme activity of neuronal and non-neuronal tissues from the freshwater snail Planorbarius corneus. The drug dose-dependently activates the basal as well as agonist-stimulated adenylyl cyclase in the ganglionic preparations, while it exerts an inhibitory effect on the enzyme present in the non-nervous tissues examined. SQ 22536 and forskolin, respectively an inhibitor and activator of adenylyl cyclase, behave as generally reported both in central and peripheral tissues of the snail. This is, to our knowledge, the first report of a stimulatory action of MDL 12330A on an adenylyl cyclase system.

Antagonizing effect of morphine on the mobility and phagocytic activity of invertebrate immunocytes.

In the present study we have demonstrated that lipopolysaccharide (LPS) acts as an activator in the immunocytes of molluscs Planorbarius corneus, Mytilus edulis, and the insect Leucophaea maderae. This stimulatory effect, demonstrated by cellular conformational changes, is concentration- and time-dependent, and is antagonized by morphine. The inhibitory effect of morphine can be counteracted by naloxone. Morphine inhibitory action on immunocyte activity is also demonstrated by a decrease in the phagocytic activity. These data suggest that the downregulation of morphine is not limited to vertebrates but is also present in invertebrates.

Microglia in invertebrate ganglia.

The results of this study lend strong support to the concept of the existence in insects and molluscs of a distinctive class of neuroglial cells comparable to vertebrate microglia. The evidence presented is as valid as that used in reference to the separate status of vertebrate microglia--i.e., the demonstration of a close structural and functional relationship of these cells with cells of the immune system. As in vertebrates, the excision of ganglia from three invertebrate species (the molluscs Planorbarius corneus and Mytilus edulis and the insect Leucophaea maderae) and their maintenance in incubation media led to an exodus of small cells and their accumulation in the culture dish. During this process, they underwent conformational changes from stellate to rounded, and then to more or less ameboid, comparable to those indicative of the process of activation in the animals' immunocytes. Functional characteristics which these translocated microglia-like cells share with immunocytes are motility, phagocytotic activity, and adherence to the culture dish. Furthermore, the two cells have certain biochemical features in common--e.g., the presence of certain cytokines and (at least in Planorbarius) that of corticotropin. An additional phenomenon of particular interest for the classification of microglial elements is their response to morphine. At 10(-6) M, this drug decreases not only the number of cells emerging from the excised ganglia but also the degree of their transformation to the "active" ameboid form. This dose-dependent and naloxone-sensitive effect of morphine on microglial cells parallels that on activated immunocytes of the same species. Corresponding results demonstrating an inhibitory effect of morphine on mobilized microglial cells of the frog Rana pipiens indicate that this relationship between the two cell types under consideration also exists in vertebrates. Binding and displacement experiments with membrane homogenates of microglial cells as well as immunocytes of Mytilus have shown that the effects of morphine on both cell types are mediated by the same special opiate receptor (mu 3).

Effects of the small cardioactive peptide B (SCPB) on adenylate cyclase of the central nervous system and peripheral organs of the freshwater snail Planorbarius corneus.

The effect of the small cardioactive peptide B (SCPB) on adenylate cyclase activity of the central nervous system (CNS) and some peripheral organs of the freshwater snail Planorbarius corneus was investigated. This peptide stimulates enzyme activity in a dose-dependent manner in all the tested ganglia, in the salivary glands, the buccal mass and the oesophagus. The amount required for half-maximal stimulation is approx 0.1 microM. Moreover, the response of adenylate cyclase to SCPB progressively increases with time, without desensitizing. On the other hand, the peptide is ineffective on heart adenylate cyclase. The responsiveness of adenylate cyclase activity to the peptide and the wide distribution of SCPB-related immunoreactivity in all the tested ganglia suggest that this substance may exert an important role as neurotransmitter as well as neuromodulator in the CNS of the snail. Since SCPB is consistently present in the buccal and cerebral ganglia, associated nerves, salivary glands, buccal mass and oesophagus, and in the light of the appreciable effects exerted by SCPB on adenylate cyclase activity in the examined peripheral tissues, we suggest that the peptide is strongly involved in the control of feeding behaviour of P. corneus.

Cell-specific effects of lead on cultured neurons of the freshwater snail Planorbarius corneus.

Cultures of isolated neuronal populations from the central ganglia of the gastropod mollusc Planorbarius corneus were used for testing the effects of inorganic lead. The examined parameters were cell survival, neurite outgrowth and cytoskeletal morphology. In large heterogeneous neuronal populations as obtained from a whole cerebral or pedal ganglion, the different sensitivity to lead is reflected mainly on the cell survival. The neurons belonging to the homogeneous E cluster population are more sensitive; in fact a higher percentage of them do not survive in the presence of lead. Moreover, in this neuronal cell type the neurite outgrowth is dramatically affected by lead only when the neurons are cultured on conditioned substrate. Possibly, membrane mechanisms activated for the neurite outgrowth represent a target for inorganic lead. The few neurites sprouted in presence of lead do not evidence changes in the cytoskeletal components.

Occurrence and distribution of insulin receptor-like immunoreactivity in molluscan brains.

Insulin receptor-like immunoreactivity (IR) was investigated in the central ganglia of different gastropod molluscs using a monoclonal antiserum raised against an epitope of the human placental insulin receptor. A well detectable and clear location of immunoreactive material was mainly found in the CNS of the freshwater snails Planorbarius corneus and Lymnaea stagnalis. This evidence correlates quite suggestively with the occurrence in these snails of a molluscan insulin peptide (MIP) previously shown. The widespread and discrete distribution of IR over a variety of physiologically distinct regions of P. corneus and L. stagnalis ganglia argues that these "receptors" may modulate multiple functions within the CNS.

Light- and electron-microscopic immunocytochemistry of a molluscan insulin-related peptide in the central nervous system of Planorbarius corneus.

Two groups of cerebral dorsal cells of the pulmonate snail Planorbarius corneus stain positively with antisera raised against synthetic fragments of the B- and C-chain of the molluscan pro-insulin-related prohormone, proMIP-I, of another pulmonate snail, Lymnaea stagnalis. At the light-microscopic level the somata of the dorsal cells and their axons and neurohemal axon terminals in the periphery of the paired median lip nerves are immunoreactive with both antisera. Furthermore, the canopy cells in the lateral lobes of the cerebral ganglia are positive. In addition, MIPB-immunoreactive neurons are found in most other ganglia of the central nervous system. At the ultrastructural level, pale and dark secretory granules are found in somata and axon terminals of the dorsal cells. Dark granules are about 4 times as immunoreactive to both antisera as pale granules. Release of anti-MIPB- and anti-MIPC-immunopositive contents of the secretory granules by exocytosis is apparent in material treated according to the tannic acid method. It is concluded that the dorsal and canopy cells synthesize a molluscan insulin-related peptide that is packed in the cell body into secretory granules and that is subsequently transported to the neurohemal axon terminals and released into the hemolymph by exocytosis. Thus, MIP seems to act as a neurhormone on peripheral targets. On the basis of the analogy between the dorsal cells and the MIP-producing cells in L. stagnalis, it is proposed that the dorsal cells of P. corneus are involved in the control of body growth and associated processes.

Amplification of GC-rich DNAs in neuronal nuclei of Planorbarius corneus (L.) (Mollusca, pulmonata).

Quantitative microfluorometric evaluation of DNA content in nerve cells of the Pulmonate Gastropod Planorbarius corneus has indicated that the increase in nuclear volume is due to DNA amplification. Indeed, it has been observed that the DNA contents are scattered at random between 2C and 1,000C values. This is not in agreement with the occurrence of repeated duplications of the whole genome. Furthermore, chromatin photo-oxidation, a technique useful in discriminating GC-rich from AT-rich DNAs, suggests that DNA amplification involves GC-rich sequences.

Effects of serotonin, dopamine and prostaglandin E(2) on adenylate cyclase activity and cyclic AMP levels in different ganglia of the freshwater snail Planorbis corneus L.

The effects of different neuroactive agents on cyclic AMP level of selected ganglia of Planorbis corneus were studied. Serotonin, dopamine and prostaglandin E(2) were capable of increasing significantly cyclic AMP synthesis in all the preparations. When such substances were tested in pairs, supra-additive effects were always observed. In high Ca(2+)-high Mg(2+) solutions dopamine action was blocked, meanwhile serotonin and prostaglandin E(2) were still effective in stimulating cyclic AMP synthesis. In the same experimental condition the supra-additive increases of the nucleotide level by drug combinations disappeared. Serotonin, but not dopamine, significantly stimulated adenylate cyclase activity in all the preparations, while prostaglandin E(2) was effective only in the Viscero-Parietal Complex. The presence of the adenylate cyclase activity in the nervous tissue of Planaorbis was substained by histochemical studies. These results demonstrating that in the nervous system of Planorbis cyclic AMP level is affected by neurotransmitters and neuromodulators, might support the idea of the crucial role of the cyclic nuclotide in the modulation of synaptic transmission.