Accumulation and decomposition of cultural residues of Theobroma grandiflorum, Paullinia cupana, Bixa orellana and forest in the southern region of Amazonas.

The litter deposited on the soil surface at various stages of decomposition is important for primary productivity that impacts the microbial communities and soil carbon storage. The objective of this study was to evaluate the accumulation and decomposition of cultural residues of Theobroma grandiflorum (Willd. ex. Spreng) Schum, Paullinia cupana (Mart.) Ducke, Bixa orellana L., and forest in the Amazon region. The study was carried out in the São Francisco settlement, Canutama in the south of Amazonas, in a randomized block experimental design, and the treatments consisted of four areas with different crops: 1 - P. cupana; 2 - T. grandiflorum; 3 - B. orellana; 4 - Native woodland area (forest), in time subdivided plots: 7, 15, 30, 60, 90, 120, 150, 180, 210, 240, 270, 300, and 330 days after the distribution of the bags in the field, all with four repetitions. To evaluate the contribution and fractions of litter, conical collectors were used in each area, and collections were performed monthly in the period from March 2020 to February 2021. The estimate of the decomposition rate of the litter was done by quantifying the loss of mass, using litter bags, which allow for a direct analysis of the rate of decay over time. The forest and P. cupana environments presented the highest litter production, and greater deposition when compared to environments cultivated with T. grandiflorum and B. orellana. The forest and B. orellana areas showed the highest speed of decomposition, while the opposite situation occurred under T. grandiflorum and P. cupana cultivation.

Muon implantation experiments in films: Obtaining depth-resolved information.

Implanted positive muons with low energies (in the range 1-30 keV) are extremely useful local probes in the study of thin films and multi-layer structures. The average muon stopping depth, typically in the order of tens of nanometers, is a function of the muon implantation energy and of the density of the material, but the stopping range extends over a broad region, which is also in the order of tens of nanometers. Therefore, an adequate simulation procedure is required in order to extract the depth dependence of the experimental parameters. Here, we present a method to extract depth-resolved information from the implantation energy dependence of the experimental parameters in a low-energy muon spin spectroscopy experiment. The method and corresponding results are exemplified for a semiconductor film, Cu(In,Ga)Se2, covered with a thin layer of Al2O3, but can be applied to any heterostructure studied with low-energy muons. It is shown that if an effect is present in the experimental data, this method is an important tool to identify its location and depth extent.

Acute antinociceptive effect of fish oil or its major compounds, eicosapentaenoic and docosahexaenoic acids on diabetic neuropathic pain depends on opioid system activation.

Diabetic neuropathic pain is one of the most common and debilitating complications of diabetes whose available treatments are poorly effective. Currently, omega-3 polyunsaturated fatty acids (ω-3 PUFAs) have been widely studied as a treatment of many types of pain, including inflammatory, spontaneous and neuropathic pain. However, little is known about the potential antinociceptive effect of ω-3 PUFAs (fish oil; FO or its major fatty acids, eicosapentaenoic -EPA and docosahexaenoic acids-DHA), in diabetic neuropathic pain as well as the mechanisms involved. To test, streptozotocin (STZ) -induced diabetic male Wistar rats were submitted to acute treatment with FO, EPA or DHA at the second and fourth weeks after diabetes induction (at the beginning and peak of development of mechanical allodynia, respectively). The cumulative effect of these compounds after a sub-chronic treatment for two weeks was also evaluated as well as the role of central µ-opioid receptors. It was observed that acute oral treatment with FO (0.5, 1 or 3 g/kg), EPA or DHA (100, 200 or 400 mg/kg) at the 2nd or at the 4th week after STZ significantly reverted the mechanical allodynia of diabetic animals, without altering the hyperglycemia or reduced weight gain. Moreover, the sub-chronic treatment with FO, EPA or DHA induced a sustained antinociceptive effect in diabetic animals. Intriguingly, the intrathecal treatment with a µ-opioid receptor antagonist (CTOP; 10 µg/rat) completely prevented the acute effect of FO, EPA or DHA. Taken together, our data suggest that ω-3 PUFAs may represent a promising therapeutic outcome for diabetic neuropathic pain, probably acting through the opioid system activation.

Dactylogyrids (Platyhelminthes, Monogenoidea) from the gills of Hoplias malabaricus (Characiformes: Erythrinidae) from coastal rivers of the Oriental Amazon Basin: species of Urocleidoides and Constrictoanchoratus n. gen.

Five species of Urocleidoides (one new) and two new species of Constrictoanchoratus n. gen. are described in this study. All were collected from the gills of Hoplias malabaricus (Characiformes: Erythrinidae) captured in six localities of coastal rivers of the north-eastern sector the State of Pará (Oriental Amazon): Urocleidoides brasiliensis Rosim, Mendoza-Franco & Luque, 2011; Urocleidoides bulbophallus n. sp.; Urocleidoides cuiabai Rosim, Mendoza-Franco & Luque, 2011; Urocleidoides eremitus Kritsky, Thatcher & Boeger, 1986; Urocleidoides malabaricusi Rosim, Mendoza-Franco & Luque, 2011; Constrictoanchoratus lemmyi n. gen. n. sp.; and Constrictoanchoratus ptilonophallus n. gen. n. sp. This is the first reported occurrence of the four previously described species of Urocleidoides parasitizing H. malabaricus from streams in the Oriental Amazon Basin. The analysis of voucher specimens of U. eremitus parasitizing the gills of H. malabaricus from the Upper Paraná River floodplain in the limits of States of Paraná and Mato Grosso do Sul, Brazil, indicates that these specimens are members of a new species of Urocleidoides, described here as Urocleidoides paranae n. sp. Constrictoanchoratus n. gen. is proposed for the species with a male copulatory organ sclerotized, coiled, clockwise; ventral anchor with elongate superficial root, inconspicuous deep root; dorsal anchor with inconspicuous roots, and a constriction at the intersection between the shaft and the point. The host-parasite diversity scenario and host specificity of the species of Constrictoanchoratus n. gen. and Urocleidoides from the gills of H. malabaricus are also discussed in this study.

Involvement of sulfated glycosaminoglycans on the development and attachment of Trypanosoma cruzi to the luminal midgut surface in the vector, Rhodnius prolixus.

In the present study, we investigated the involvement of sulfated glycosaminoglycans in both the in vivo development and adhesion of T. cruzi epimastigotes to the luminal surface of the digestive tract of the insect vector, Rhodnius prolixus. Pre-incubation of T. cruzi, Dm 28c epimastigotes with heparin, chondroitin 4-sulfate, chondroitin 6-sulfate or protamine chloridrate inhibited in vitro attachment of parasites to the insect midgut. Enzymatic removal of heparan sulfate moieties by heparinase I or of chondroitin sulfate moieties by chondroitinase AC from the insect posterior midgut abolished epimastigote attachment in vitro. These treatments also reduced the labelling of anionic sites exposed at the luminal surface of the perimicrovillar membranes in the triatomine midgut epithelial cells. Inclusion of chondroitin 4-sulfate or chondroitin 6-sulfate and to a lesser extent, heparin, in the T. cruzi-infected bloodmeal inhibited the establishment of parasites in R. prolixus. These observations indicate that sulfated glycosaminoglycans are one of the determinants for both adhesion of the T. cruzi epimastigotes to the posterior midgut epithelial cells of the triatomine and the parasite infection in the insect vector, R. prolixus.

Streptozotocin-induced mechanical hypernociception is not dependent on hyperglycemia.

Since streptozotocin (STZ)-induced diabetes is a widely used model of painful diabetic neuropathy, the aim of the present study was to design a rational protocol to investigate whether the development of mechanical hypernociception induced by STZ depends exclusively on hyperglycemia. Male Wistar rats (180-200 g; N = 6-7 per group) received a single intravenous injection of STZ at three different doses (10, 20, or 40 mg/kg). Only the higher dose (40 mg/kg) induced a significant increase in blood glucose levels, glucose tolerance and deficiency in weight gain. However, all STZ-treated rats (hyperglycemic or not) developed persistent (for at least 20 days) and indistinguishable bilateral mechanical hypernociception that was not prevented by daily insulin treatment (2 IU twice a day, sc). Systemic morphine (2 mg/kg) but not local (intraplantar) morphine treatment (8 microg/paw) significantly inhibited the mechanical hypernociception induced by STZ (10 or 40 mg/kg). In addition, intraplantar injection of STZ at doses that did not cause hyperglycemia (30, 100 or 300 microg/paw) induced ipsilateral mechanical hypernociception for at least 8 h that was inhibited by local and systemic morphine treatment (8 microg/paw or 2 mg/kg, respectively), but not by dexamethasone (1 mg/kg, sc). The results of this study demonstrate that systemic administration of STZ induces mechanical hypernociception that does not depend on hyperglycemia and intraplantar STZ induces mechanical sensitization of primary sensory neurons responsive to local morphine treatment.

Streptozotocin-induced mechanical hypernociception is not dependent on hyperglycemia

Since streptozotocin (STZ)-induced diabetes is a widely used model of painful diabetic neuropathy, the aim of the present study was to design a rational protocol to investigate whether the development of mechanical hypernociception induced by STZ depends exclusively on hyperglycemia. Male Wistar rats (180-200 g; N = 6-7 per group) received a single intravenous injection of STZ at three different doses (10, 20, or 40 mg/kg). Only the higher dose (40 mg/kg) induced a significant increase in blood glucose levels, glucose tolerance and deficiency in weight gain. However, all STZ-treated rats (hyperglycemic or not) developed persistent (for at least 20 days) and indistinguishable bilateral mechanical hypernociception that was not prevented by daily insulin treatment (2 IU twice a day, sc). Systemic morphine (2 mg/kg) but not local (intraplantar) morphine treatment (8 µg/paw) significantly inhibited the mechanical hypernociception induced by STZ (10 or 40 mg/kg). In addition, intraplantar injection of STZ at doses that did not cause hyperglycemia (30, 100 or 300 µg/paw) induced ipsilateral mechanical hypernociception for at least 8 h that was inhibited by local and systemic morphine treatment (8 µg/paw or 2 mg/kg, respectively), but not by dexamethasone (1 mg/kg, sc). The results of this study demonstrate that systemic administration of STZ induces mechanical hypernociception that does not depend on hyperglycemia and intraplantar STZ induces mechanical sensitization of primary sensory neurons responsive to local morphine treatment.

Molecular characterization of patients with X-linked Hyper-IgM syndrome: description of two novel CD40L mutations.

Type 1, X-linked Hyper-IgM syndrome (HIGM1) is caused by mutations in the gene encoding the CD154 protein, also known as CD40 ligand (CD40LG). CD40L is expressed in activated T cells and interacts with CD40 receptor expressed on B lymphocytes and dendritic cells. Affected patients present cellular and humoral immune defects, with infections by intracellular, opportunistic and extracellular pathogens. In the present study we investigated the molecular defects underlying disease in four patients with HIGM1. We identified four distinct CD40L mutations, two of them which have not been previously described. P1 harboured the novel p.G227X mutation which abolished CD40L expression. P2 had a previously described frame shift deletion in exon 2 (p.I53fsX65) which also prevented protein expression. P3 demonstrated the previously known p.V126D change in exon 4, affecting the TNF homology (TNFH) domain. Finally, P4 evidenced the novel p.F229L mutation also located in the TNFH domain. In silico analysis of F229L predicted the change to be pathological, affecting the many hydrophobic interactions of this residue. Precise molecular diagnosis in HIGM syndrome allows reliable detection of carriers, making genetic counselling and prenatal diagnosis possible.

Trypanosoma cruzi: attachment to perimicrovillar membrane glycoproteins of Rhodnius prolixus.

Studies were carried out to identify proteins involved in the interface of Trypanosoma cruzi with the perimicrovillar membranes (PMM) of Rhodnius prolixus. Video microscopy experiments demonstrated high level of adhesion of T. cruzi Dm 28c epimastigotes to the surface of posterior midgut cells of non-treated R. prolixus. The parasites however were unable to attach to gut cells obtained from decapitated or azadirachtin-treated insects. The influence of carbohydrates on the adhesion to insect midgut was confirmed by inhibition of parasite attachment after midgut incubation with N-acetylgalactosamine, N-acetylmannosamine, N-acetylglucosamine, D-galactose, D-mannose or sialic acid. We observed that hydrophobic proteins in the surface of epimastigotes bind to polypeptides with 47.7, 45.5, 44, 43, 40.5, 36, 31 and 13kDa from R. prolixus PMM and that pre-incubation of lectins specifically inhibited binding to 31, 40.5, 44 and 45.5kDa proteins. We suggest that glycoproteins from PMM and hydrophobic proteins from epimastigotes are important for the adhesion of the parasite to the posterior midgut cells of the vector.

Antiserum against perimicrovillar membranes and midgut tissue reduces the development of Trypanosoma cruzi in the insect vector, Rhodnius prolixus.

Antiserum raised against Rhodnius prolixus perimicrovillar membranes (PMM) and midgut tissue interfered with the midgut structural organization and reduced the development of Trypanosoma cruzi in the R. prolixus insect vector. SDS-PAGE and Western blot analyses confirmed the specific recognition of midgut proteins by the antibody. Feeding, mortality, molt, and oviposition of the insects were unaffected by feeding with the antiserum. However, the eclosion of the eggs were reduced from R. prolixus females treated with antiserum. Additionally, in vivo evaluation showed that after oral treatment with the antiserum, the intensity of infection with the Dm-28c clone of T. cruzi decreased in the digestive tract of fifth-instar nymphs and in the excretions of R. prolixus adults. These results suggest that the changes observed in the PMM organization in the posterior midgut of R. prolixus may not be important for triatomine survival but the antiserum acts as a transmission-reduction vaccine able to induce significant decreases in T. cruzi infection in the vector.

Effect of blood components, abdominal distension, and ecdysone therapy on the ultrastructural organization of posterior midgut epithelial cells and perimicrovillar membranes in Rhodnius prolixus

The effects of blood components, nerve-cord severance, and ecdysone therapy on the posterior midgut epithelial cells of 5th-instar Rhodnius prolixus nymphs 10 days after feeding were analyzed by transmission electron microscopy. Cutting the nerve-cord of the blood-fed insects partially reduced the development of microvilli and perimicrovillar membranes (PMM), and produced large vacuoles and small electrondense granules; insects fed on Ringer's saline diet exhibited well developed microvilli and low PMM production; swolled rough endoplasmatic reticulum and electrondense granules; Ringer's saline meal with ecdysone led to PMM development, glycogen particles, and several mitochondria in the cytoplasm; epithelial cells of the insects fed on Ringer's saline meal whose nerve-cord was severed showed heterogeneously distributed microvilli with reduced PMM production and a great quantity of mitochondria and glycogen in the cytoplasm; well developed microvilli and PMM were observed in nerve-cord severed insects fed on Ringer's saline meal with ecdysone; Ringer's saline diet containing hemoglobin recovered the release of PMM; and insects fed on human plasma showed slightly reduced PMM production, although the addition of ecdysone in the plasma led to a normal midgut ultrastructural organization. We suggest that the full development of microvilli and PMM in the epithelial cells depends on the abdominal distension in addition to ingestion of hemoglobin, and the release of ecdysone.

Effect of blood components, abdominal distension, and ecdysone therapy on the ultrastructural organization of posterior midgut epithelial cells and perimicrovillar membranes in Rhodnius prolixus.

The effects of blood components, nerve-cord severance, and ecdysone therapy on the posterior midgut epithelial cells of 5th-instar Rhodnius prolixus nymphs 10 days after feeding were analyzed by transmission electron microscopy. Cutting the nerve-cord of the blood-fed insects partially reduced the development of microvilli and perimicrovillar membranes (PMM), and produced large vacuoles and small electrondense granules; insects fed on Ringer's saline diet exhibited well developed microvilli and low PMM production; swolled rough endoplasmatic reticulum and electrondense granules; Ringer's saline meal with ecdysone led to PMM development, glycogen particles, and several mitochondria in the cytoplasm; epithelial cells of the insects fed on Ringer's saline meal whose nerve-cord was severed showed heterogeneously distributed microvilli with reduced PMM production and a great quantity of mitochondria and glycogen in the cytoplasm; well developed microvilli and PMM were observed in nerve-cord severed insects fed on Ringer's saline meal with ecdysone; Ringer's saline diet containing hemoglobin recovered the release of PMM; and insects fed on human plasma showed slightly reduced PMM production, although the addition of ecdysone in the plasma led to a normal midgut ultrastructural organization. We suggest that the full development of microvilli and PMM in the epithelial cells depends on the abdominal distension in addition to ingestion of hemoglobin, and the release of ecdysone.

Cytokine-mediated inflammatory hyperalgesia limited by interleukin-1 receptor antagonist.

1. The effect of IL-1ra on response to intraplantar (i.pl.) injection of LPS, carrageenin, bradykinin, TNFalpha, IL-1beta, IL-8, PGE(2) and dopamine was investigated in a model of mechanical hyperalgesia in rats. 2. IL-1ra inhibited hyperalgesic response to LPS, carrageenin, bradykinin, TNFalpha, and IL-1beta, but not responses to IL-8, PGE(2) and dopamine. 3. A sheep anti-rat IL-1ra serum potentiated response to LPS, carrageenin, bradykinin, TNFalpha and IL-1beta but not IL-8. 4. Carrageenin and LPS stimulated and production of immunoreactive TNFalpha, IL-1beta and IL-1ra in the skin of injected paws. 5. The inhibition by IL-1ra of the hyperalgesic response to carrageenin was not affected by antibodies neutralizing IL-4 and IL-10. 6. In mice, IL-1ra inhibited the nociceptive response to i.p. injection of acetic acid. 7. These data suggest that IL-1ra, released at sites of inflammation, limits inflammatory hyperalgesia. This effect is independent of (IL-1ra-induced) IL-4 and IL-10 and appears to be the result of antagonism by IL-1ra of IL-1beta-stimulated eicosanoid production.

Bradykinin B1 and B2 receptors, tumour necrosis factor alpha and inflammatory hyperalgesia.

The effects of BK agonists and antagonists, and other hyperalgesic/antihyperalgesic drugs were measured (3 h after injection of hyperalgesic drugs) in a model of mechanical hyperalgesia (the end-point of which was indicated by a brief apnoea, the retraction of the head and forepaws, and muscular tremor). DALBK inhibited responses to carrageenin, bradykinin, DABK, and kallidin. Responses to kallidin and DABK were inhibited by indomethacin or atenolol and abolished by the combination of indomethacin + atenolol. DALBK or HOE 140, given 30 min before, but not 2 h after, carrageenin, BK, DABK and kallidin reduced hyperalgesic responses to these agents. A small dose of DABK+ a small dose of BK evoked a response similar to the response to a much larger dose of DABK or BK, given alone. Responses to BK were antagonized by HOE 140 whereas DALBK antagonized only responses to larger doses of BK. The combination of a small dose of DALBK with a small dose of HOE 140 abolished the response to BK. The hyperalgesic response to LPS (1 microg) was inhibited by DALBK or HOE 140 and abolished by DALBK + HOE 140. The hyperalgesic response to LPS (5 microg) was not antagonized by DALBK + HOE 140. These data suggest: (a) a predominant role for B2 receptors in mediating hyperalgesic responses to BK and to drugs that stimulate BK release, and (b) activation of the hyperalgesic cytokine cascade independently of both B1 and B2 receptors if the hyperalgesic stimulus is of sufficient magnitude.

Trypanosoma cruzi: both chemically induced and triatomine-derived metacyclic trypomastigotes cause the same immunological disturbances in the infected mammalian host.

Infection of BALB/c mice with chemically induced metacyclic forms of Trypanosoma cruzi clone Dm28c led to characteristic changes of experimental Chagas' disease, with protracted but marked parasitemia, intense splenomegaly, and splenic T cell hyporeactivity to TcR;CD3-dependent stimulation. Infection of BALB/c mice with either chemically induced or triatomine-derived Dm28c metacyclic forms led to comparable parasitemias, a synchronous increase in the number of splenic large lymphocytes, and a similar reduction in T cell responsivity to immobile anti-CD3 antibody. A marked and selective reduction in the level of CD8 expression per cell was also seen in mice infected with either form of metacyclic parasites. Large inflammatory mononuclear cell infiltrates were present in the hearts of mice infected with either chemically induced or insect vector-derived metacyclic forms, at both acute and chronic stage, with predominance of CD8 over CD4 T cells in the lesions, in both cases. These results indicate that infection with chemically induced metacyclic forms of T. cruzi can be a useful model of Chagas' disease, resembling infection caused by the insect vector.

Cyclophosphamide blocks both antigen-specific and polyclonal immunoglobulin responses in experimental visceral leishmaniasis.

Cyclophosphamide (Cy) has been shown to modulate antibody responses in a wide range of diseases both in humans and experimental animals. Our results in Syrian hamsters infected with Leishmania donovani have shown that Cy blocks specific and polyclonal antibody production both in vivo and in vitro. This effect was achieved by weekly 100 mg/kg doses and also by a 300 mg/kg single dose. Although Cy provokes a significant decrease in B-cell numbers in infected animals, this cannot explain the suppression of antibody production since a 50% decrease in B-cells of only-infected hamsters did not reproduce the same effect in in vitro assays. Also, this suppression was not reversed either by elimination of adherent cells or by the presence of indomethacin. These data suggest that Cy affects T-cell populations involved in the control of antibody production by B-cells.

[Characteristics of intra-metropolitan mobility in the state of Sao Paulo, Brazil, 1970-1980]. / Caracteristicas de la movilidad intrametropolitana en el estado de Sao Paulo, Brasil, 1970-1980.

"Based on the 1980 census, the study attempts to provide a general overview of [intra-metropolitan migration in Brazil], using information regarding previous place of residence--data included for the first time in a Brazilian census--, which helps to establish the internal, intermunicipal, migratory flow. The article also presents certain aspects related to volume, direction and reasons for these movements as well as some of the characteristics of influencing factors." (SUMMARY IN ENG)

Cyclophosphamide affects the dynamics of granuloma formation in experimental visceral leishmaniasis.

We observed histopathological and ultrastructural hepatic changes following the intracardiac inoculation of Leishmania donovani amastigotes into inbred LHC hamsters (group I). Since granuloma formation is known to be T-cell-dependent, we also examined infected hamsters under cyclophosphamide immunosuppressive treatment (group ICy) and evaluated the production of interleukin-2 (IL-2) by their cells. Group I showed more intense hepatocyte and endothelial cell clasmatosis as well as hepatocyte degeneration and necrosis, deposits of connective tissue fibers, granulomas with multinucleated giant cells (MGCs) of foreign-body and Langhans' types and reduced production of IL-2 by spleen cells. In contrast, group ICy hamsters exhibited larger eosinophil and lymphocyte populations within sinusoids and peri-sinusoidal areas but showed no MGCs in granulomas. A striking decline in IL-2 production was noted. These results suggest that cyclophosphamide induces a delay in the natural evolution of L. donovani-induced granulomatous hepatic inflammation.

Jejunal biopsy in HIV-infected patients.

Forty-nine HIV-infected patients were submitted to peroral jejunal biopsy in order to evaluate the presence of microorganisms and the histomorphometric aspects of the enteric mucosa with subsequent correlation of these findings to the appropriate clinical stage of the disease. Thirty-seven patients fulfilled the CDC criteria for AIDS, of whom 23 presented with diarrhea. Of the 12 patients who had not yet been given an AIDS diagnosis. 3 had persistent generalized lymphadenopathy and 9 were asymptomatic carriers. Flat mucosa was observed in two patients (8.7%) with diarrhea and coccidea. Subtotal villous atrophy and severe lamina propria (LP) mononuclear infiltrate (13%) were found only in patients with diarrhea. Moderate to severe histologic changes were more frequently observed in this group, not always related to the presence of microorganisms. Crypt hyperregeneration was a constant finding. Intraepithelial lymphocyte (IEL) count was decreased in patients with diarrhea. Specific infectious agents were unexpectedly rare for the tropical developing country population studied. The organism most commonly associated with diarrhea was Cryptosporidium sp. (21.7%). The etiology of diarrhea in a significant number of patients remains unclear.

Effects of cannabidiol in animal models predictive of antipsychotic activity.

The effects of cannabidiol (CBD) were compared to those produced by haloperidol in rats submitted to experimental models predictive of antipsychotic activity. Several doses of CBD (15-480 mg/kg) and haloperidol (0.062-1.0 mg/kg) were tested in each model. First, CBD increased the effective doses 50% (or) ED50 of apomorphine for induction of the sniffing and biting stereotyped behaviors. In addition, both CBD and haloperidol reduced the occurrence of stereotyped biting induced by apomorphine (6.4 mg/kg), increased plasma prolactin levels and produced palpebral ptosis, as compared to control solutions. However, CBD did not induce catalepsy even at the highest doses, in contrast to haloperidol. Such a pharmacological profile is compatible with that of an "atypical" antipsychotic agent, though the mechanism of action is uncertain and may not be identical to that of the dopamine antagonists.