Spontaneous and precipitated withdrawal with a synthetic cannabinoid, WIN 55212-2.

Physical dependence on the synthetic cannabinoid-receptor agonist R(+)-[2,3-dihydro-5-methyl-3-[(morpholinyl)methyl]pyrrolo[1,2,3-de]-1,4-benzoxazinyl]-(1-naphthalenyl) methanone mesylate (WIN 55212-2) was demonstrated in rats by the use of a chronic continuous infusion. Spontaneous withdrawal, of moderate intensity, was shown for the first time with this class of drugs of abuse. Behavioral withdrawal signs were also elicited after challenge with (N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide.HCl (SR141716A), a specific CB(1) cannabinoid-receptor antagonist. In both instances, the high-dose regimen (4, 8, 16 and 16 mg/kg/day, i.p. on days 1-4, respectively) was sufficient to evoke a typical withdrawal syndrome quantified by the signs wet-dog shakes and facial rubs. These results are discussed relative to those obtained with Delta(9)-tetrahydrocannabinol and anandamide. With Delta(9)-tetrahydrocannabinol, precipitated but not spontaneous or abrupt withdrawal was observed, and this was ascribed to pharmacokinetic properties. Anandamide, which showed little, if any, physical dependence potential, behaved atypically. Possible implications regarding pharmacotherapeutic and human abuse issues are discussed.

Dihydroetorphine: physical dependence and stereotypy after continuous infusion in the rat.

In a previous study in this laboratory, exposure of rhesus monkeys to intermittent, high doses of dihydroetorphine for 42 days did not evoke behavioral signs of physical dependence on this opioid either after it was abruptly withdrawn or after challenge with a high dose of naloxone. To investigate further the physical dependence capacity of this opioid, it was given by infusion to rats thereby exposing receptors chronically and continuously to this opioid. Abstinence expressed as body weight loss, irritability, and wet-dog shakes was observed after abrupt withdrawal of the low-dose regimen (5,10, 40 and 40 microg/kg per day for 4 days, respectively). The high-dose regimen (10, 20 and 80 microg/kg per day for 3 days, respectively) produced stereotypy and physical dependence. Although many reported molecular events and dependence studies suggest otherwise, dihydroetorphine's propensity to produce physical dependence, an important determinant of opioid abuse, is real.

Anandamide, an endogenous cannabinoid, has a very low physical dependence potential.

Using N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2, 4-dichloro-phenyl)-4-methyl-1H-pyrazole-3-carboxamide. HCl (SR 141716A), a cannabinoid antagonist, several investigators (deFonseca et al., 1997; Aceto et al., 1995, 1996; Tsou et al., 1995) demonstrated physical dependence on THC [Delta9-tetrahydrocannabinol]. This demonstration prompted us to determine whether anandamide, an endogenous cannabinoid agonist, would also produce physical dependence. A low-dose regimen (10, 20, 40 and 40) or a high-dose regimen (25, 50, 100 and 100) expressed as mg/kg/24 hr was infused i.p. on a continuous basis, from days 1 through 4, respectively. During the infusion, especially at the high-dose regimen, the rats became immobile and developed eyelid ptosis. Abrupt discontinuation of anandamide did not elicit rebound behavioral activity. Neither arachidonic acid, a precursor and metabolite of anandamide (50, 100, 200 and 200 mg/kg/24 hr on days 1 through 4, respectively), nor 2-Me-F-AN [2-methylarachidonyl-(2'-fluoroethyl)-amide], a metabolically stable analog of anandamide (5, 10, 20 and 20 mg/kg/24 hr for 4 days, respectively), had remarkable effects. Notably, groups pretreated with anandamide or 2-Me-F-AN and challenged with SR 141716A did not show significantly elevated behavioral scores when compared with SR 141716A controls. On the other hand, nearly all groups receiving SR 141716A showed significant activation of these behaviors compared with vehicle controls, which suggests that this cannabinoid antagonist itself was activating behavior. We concluded that anandamide has little if any capacity for physical dependence. The finding that SR 141716A activated behavior supports the hypothesis that the cannabimimetic system exerts a depressant effect in the CNS.

Dependence on delta 9-tetrahydrocannabinol: studies on precipitated and abrupt withdrawal.

A cannabinoid antagonist, SR 141716A, dose dependently precipitated a behavioral withdrawal syndrome in rats continuously infused i.p. for only 4 days with relatively low-dose regimens of delta 9-tetrahydrocannabinol. The following dose regimens, expressed as mg/kg/24 hr, were used for days 1 through 4: high-12.5, 25, 50 and 100; medium-2.5, 5, 10 and 20; and low-0.5, 1, 2 and 4. The major withdrawal signs of the syndrome were scratching, rubbing face with paws, licking, wetdog shakes, arched back and ptosis (at least 50% closure of eyelids). At the highest dose regimen, other signs noted in fewer subjects were biting, tongue rolling, retropulsion, head shakes, extended limbs or high stepping, ataxia, myoclonic spasms and front paw treading. During abrupt withdrawal (delta 9 tetrahydrocannabinol was discontinued and vehicle substituted) abstinence signs were also noted; however, except during a 48-hr observation period, withdrawal was not sufficiently robust to achieve statistical significance. The results of this study provide evidence that a modest course of delta 9-tetrahydrocannabinol can produce physical dependence. Hence, the risk and incidence of marijuana dependence in humans may be greater than previously projected.

Cannabinoid precipitated withdrawal by the selective cannabinoid receptor antagonist, SR 141716A.

Precipitated withdrawal in rats chronically exposed to delta 9-tetrahydrocannabinol, the major psychoactive principle of the marijuana plant, was unequivocally demonstrated for the first time using a selective antagonist, SR 141716A (N-(piperidin-1-yl)-5-(4-chlorophenyl)-1(2,4- dichloro-phenyl)-4-methyl-1H-pyrazole carboxamide.HCl). This demonstration should provide a powerful stimulus for the systematic study of dependency on the psychoactive cannabinoids.

Nicotine's opioid and anti-opioid interactions: proposed role in smoking behavior.

Nicotine produced antinociception in mice which was antagonized noncompetitively by naloxone. In addition, at significantly lower doses, nicotine noncompetitively antagonized morphine-induced antinociception. A speculative suggestion regarding the opiatergic and anti-opiatergic actions of nicotine is that it significantly promotes and maintains smoking behavior.

Diagnosis and treatment of cancer-related thrombosis.

Thrombotic events are common causes of morbidity and mortality in patients with neoplastic diseases. Standard diagnostic tests may give misleading results, and the effects of treatment may not be as expected. This article discusses the pathophysiology, diagnosis, and treatment of thrombosis in this difficult setting.

Neutropenia in an extremely premature infant treated with recombinant human granulocyte colony-stimulating factor.

Neutropenia in the newborn is often associated with sepsis, maternal hypertension, or prematurity. We describe a 654-g infant born at 30 weeks' gestation by cesarean section due to severe maternal hypertension. His course was complicated by five episodes of sepsis, including three with group B streptococcus. The results of hematologic and immunologic studies were normal except that absolute neutrophil counts were low (less than 1 x 10(9)/L) with intermittent increases during sepsis. Human recombinant granulocyte colony-stimulating factor administered subcutaneously (10 micrograms/kg per day initially) resulted in an absolute neutrophil count of greater than 30 x 10(9)/L within 2 weeks. The dosage was lowered and the absolute neutrophil counts were maintained at 8 to 12 x 10(9)/L with no further septic episodes. The human recombinant granulocyte colony-stimulating factor therapy was discontinued after 7 months, and the patient remained healthy with an absolute neutrophil count of greater than 2 x 10(9)/L. Thus, treatment with human recombinant granulocyte colony-stimulating factor may be useful as a temporary measure for neonatal neutropenia associated with sepsis. A controlled, clinical trial is warranted.

Purification and properties of a unique superoxide dismutase from Nocardia asteroides.

A unique form of superoxide dismutase was isolated and characterized from Nocardia asteroides GUH-2. This enzyme contains 1 to 2 g atoms each of Fe, Mn, and Zn per mol and exhibits spectral properties suggestive of Fe- or Mn-containing superoxide dismutases. Its Mr = 100,000, and it is composed of four subunits of equal size which are not covalently joined. The amino acid composition of the enzyme was more closely related to the Mn- or Fe-containing enzymes of Mycobacterium species and was least related to the Cu-Zn enzyme of eukaryotes. Azide at 1 and 20 mM inhibits the activity 10 and 41%, respectively, and 5 mM H2O2 inhibits 40%, but 1 or 5 mM cyanide caused trivial effect. The immunofluorescent staining, which was specific for superoxide dismutase of N. asteroides, indicated the association of this enzyme to the outer cell wall of the organism. Further, the enzyme was shown to be selectively secreted into the medium.

Response of CBA/N x DBA2/F1 mice to Nocardia asteroides.

Immunized and nonimmunized B-lymphocyte-deficient CBD2/F1 (CBA/N x DBA/2) mice were infected with Nocardia asteroides GUH-2 by different routes of inoculation. The 50% lethal dose, organ clearance, footpad response, and antibody titers were measured. It was observed that B-cell-deficient male mice were not significantly more susceptible to infection than normal female controls even though the female CBD2/F1 mice produced antinocardial antibodies while the deficient male animals did not. Preimmunized male and female mice were identical in their ability to clear N. asteroides from the adrenals, brain, kidneys, liver, lungs, and spleen. Both DBA/2 and CBD2/F1 female mice were more susceptible than their male littermates to intravenous challenge with N. asteroides GUH-2. This enhanced susceptibility of the female mice as compared to the male littermates appeared to be due to a decreased resistance to nocardial infections in the brains of the female animals. These data indicate that antibody and certain B-lymphocyte subpopulations are not essential components in host resistance to N. asteroides GUH-2 in these mice.

Role of L-forms of Nocardia caviae in the development of chronic mycetomas in normal and immunodeficient murine models.

Single-cell suspensions of Nocardia caviae 112 were injected into normal, athymic, and asplenic mice by several different routes. The 50% lethal dose values, kill curve characteristics, histological and electron microscopic properties, organ clearance patterns, and induction of L-forms during the acute and chronic phase of disease were determined in groups of mice for up to 2 years after infection. From these data we concluded the following. (i) Athymic and asplenic animals were significantly more susceptible to N. caviae than their littermate controls regardless of inoculation route. (ii) All mice were most susceptible to lethal infection after intranasal administration and least affected when the organisms were injected into the peritoneal cavity. (iii) Chronic, progressive disease leading to the formation of mycetomas occurred only in mice injected intravenously. (iv) T-cell-deficient animals were impaired in the development of typical mycetomas. (v) L-forms of N. caviae were induced within immunocompetent hosts, whereas the cell wall-less state of the bacteria was not observed in the immunodeficient animals. (vi) Two colony types of the cell wall-deficient state were isolated from infected animals. (vii) These cell wall-deficient organisms were intimately involved in the pathogenesis of disease and bacterial persistence within the host. Finally (viii), with this strain of Nocardia, cell wall-deficient organisms played a major role in the development of the characteristic bacterial granule formed within the mycetomatous lesions 6 months to 1 year after intravenous inoculation.