Disease control of acromegaly does not prevent excess mortality in the long term: results of a nationwide survey in Italy.

OBJECTIVE: This study aimed to assess the long-term outcome of patients with acromegaly. DESIGN: This is a multicenter, retrospective, observational study which extends the mean observation period of a previously reported cohort of Italian patients with acromegaly to 15 years of follow-up. METHODS: Only patients from the centers that provided information on the life status of at least 95% of their original cohorts were included. Life status information was collected either from clinical records or from the municipal registry offices. Standardized mortality ratios (SMRs) were computed comparing data with those of the general Italian population. RESULTS: A total of 811 patients were included. There were 153 deaths, with 90 expected and an SMR of 1.7 (95% CI 1.4-2.0, p < 0.001). Death occurred after a median of 15 (women) or 16 (men) years from the diagnosis, without gender differences. Mortality remained elevated in the patients with control of disease (SMR 1.3, 95% CI 1.1-1.6). In the multivariable analysis, only older age and high IGF1 concentrations at last available follow-up visit were predictors of mortality. The oncological causes of death outweighed the cardiovascular ones, bordering on statistical significance with respect to the general population. CONCLUSIONS: Mortality remains significantly high in patients with acromegaly, irrespectively of disease status, as long as the follow-up is sufficiently long with a low rate of patients lost to follow-up. Therapy strategy including radiotherapy does not have an impact on mortality. Oncological causes of death currently outweigh the cardiovascular causes.

Octreotide and pasireotide effects on medullary thyroid carcinoma (MTC) cells growth, migration and invasion.

Medullary thyroid carcinoma (MTC) is a rare neuroendocrine neoplasm of the parafollicular thyroid C cells. Although somatostatin receptors are expressed by MTCs, treatment with octreotide has shown poor efficacy, whereas recently pasireotide has demonstrated antiproliferative effects in persistent postoperative MTCs. Aim of this study was to test the effects of octreotide and pasireotide on MTC cells proliferation, cell cycle proteins expression, MAPK activation, apoptosis, calcitonin secretion, migration and invasion in TT cell line as well as in primary MTC cultured cells. Our results showed that both octreotide and pasireotide reduced TT cell proliferation (-35.2 ± 12.1%, p < 0.001, and -25.3 ± 24.8%, p < 0.05, at 10-8 M, respectively), with concomitant inhibition of ERK phosphorylation and cyclin D1 expression. This cytostatic effect was accompanied by a proapoptotic action, with an increase of caspase3/7 activity of 1.5-fold. Moreover, both octreotide and pasireotide inhibited cell migration (-50.9 ± 11.3%, p < 0.01, and -40.5 ± 17%, p < 0.05, respectively) and invasion (-61.3 ± 35.1%, p < 0.05, and -49.7 ± 18%, p < 0.01, respectively). No effect was observed on calcitonin secretion. We then tried to extend these observations to primary cultures (n = 5). Octreotide and/or pasireotide were effective in reducing cells proliferation in 3 out of 5 tumors, and to induce cell apoptosis in 1 out of 3 MTCs. Both octreotide and pasireotide were able to reduce cell migration in all MTC tested. SST2, SST3 and SST5 were expressed in all MTC, with a tendency to increased expression of SST2 in RET mutated vs wild type MTCs. In agreement, inhibition of mutated RET in TT cells reduced SST2 expression. In conclusion, we demonstrated that octreotide and pasireotide inhibited cell proliferation and invasiveness in a subset of MTC, supporting their potential use in the control of tumor growth.

The corticotrophin-releasing factor/urocortin system regulates white fat browning in mice through paracrine mechanisms.

OBJECTIVES: The corticotrophin-releasing factor (CRF)/urocortin system is expressed in the adipose tissue of mammals, but its functional role in this tissue remains unknown. METHODS: Pharmacological manipulation of the activity of CRF receptors, CRF1 and CRF2, was performed in 3T3L1 white pre-adipocytes and T37i brown pre-adipocytes during in vitro differentiation. The expression of genes of the CRF/urocortin system and of markers of white and brown adipocytes was evaluated along with mitochondrial biogenesis and cellular oxygen consumption. Metabolic evaluation of corticosterone-deficient or supplemented Crhr1-null (Crhr1(-/-)) mice and their wild-type controls was performed along with gene expression analysis carried out in white (WAT) and brown (BAT) adipose tissues. RESULTS: Peptides of the CRF/urocortin system and their cognate receptors were expressed in both pre-adipocyte cell lines. In vitro pharmacological studies showed an inhibition of the expression of the CRF2 pathway by the constitutive activity of the CRF1 pathway. Pharmacological activation of CRF2 and, to a lesser extent, inhibition of CRF1 signaling induced molecular and functional changes indicating transdifferentiation of white pre-adipocytes and differentiation of brown pre-adipocytes. Crhr1(-/-) mice showed increased expression of CRF2 and its agonist Urocortin 2 in adipocytes that was associated to brown conversion of WAT and activation of BAT. Crhr1(-/-) mice were resistant to diet-induced obesity and glucose intolerance. Restoring physiological circulating corticosterone levels abrogated molecular changes in adipocytes and the favorable phenotype of Crhr1(-/-) mice. CONCLUSIONS: Our findings suggest the importance of the CRF2 pathway in the control of adipocyte plasticity. Increased CRF2 activity in adipocytes induces browning of WAT, differentiation of BAT and is associated with a favorable metabolic phenotype in mice lacking CRF1. Circulating corticosterone represses CRF2 activity in adipocytes and may thus regulate adipocyte physiology through the modulation of the local CRF/urocortin system. Targeting CRF receptor signaling specifically in the adipose tissue may represent a novel approach to tackle obesity.

CRF2 receptor-deficiency eliminates opiate withdrawal distress without impairing stress coping.

The opiate withdrawal syndrome is a severe stressor that powerfully triggers addictive drug intake. However, no treatment yet exists that effectively relieves opiate withdrawal distress and spares stress-coping abilities. The corticotropin-releasing factor (CRF) system mediates the stress response, but its role in opiate withdrawal distress and bodily strategies aimed to cope with is unknown. CRF-like signaling is transmitted by two receptor pathways, termed CRF(1) and CRF(2). Here, we report that CRF(2) receptor-deficient (CRF(2)(-/-)) mice lack the dysphoria-like and the anhedonia-like states of opiate withdrawal. Moreover, in CRF(2)(-/-) mice opiate withdrawal does not increase the activity of brain dynorphin, CRF and periaqueductal gray circuitry, which are major substrates of opiate withdrawal distress. Nevertheless, CRF(2) receptor-deficiency does not impair brain, neuroendocrine and autonomic stress-coping responses to opiate withdrawal. The present findings point to the CRF(2) receptor pathway as a unique target to relieve opiate withdrawal distress without impairing stress-coping abilities.

Role of the corticotropin-releasing factor receptor type 2 in the control of food intake in mice: a meal pattern analysis.

The actions of corticotropin-releasing factor (CRF) and related peptides are mediated by two receptors (CRF(1) and CRF(2)). The respective role of each subtype in the control of food intake remains poorly known. In the present study, we examined the quantity and microstructure of ingestive behavior of knockout (KO) mice lacking CRF(2) receptors and their wild-type (WT) littermates. Under basal conditions, CRF(2) KO mice showed increased nocturnal food intake, evident as an increased zenith in circadian cosinor analysis of food intake. Microstructure analysis revealed that this greater food intake reflected increased meal size, rather than meal frequency, suggesting a decreased satiating value of food. Following acute restraint stress, CRF(2) KO mice showed an intact immediate anorectic response with increased latency to eat and decreased meal size. However, CRF(2) deletion abolished the prolonged phase of restraint-induced anorexia. CRF(2) KO mice did not differ from WT controls in feeding responses to food deprivation or injection of ghrelin receptor agonists. Independent of genotype, food deprivation increased food intake, with dramatic changes in meal size, meal frequency, water : food ratio and eating rate. Acyl-ghrelin or BIM-28131, a potent ghrelin analog, dose-dependently stimulated food intake by increasing meal size (ghrelin, BIM-28131) and meal number (BIM-28131), while slowing the average eating rate (BIM-28131) similarly in WT and KO mice. These results suggest that the CRF(2) receptor is involved in the control of meal size during the active phase of eating and following acute exposure to stress.

Targeted mutations of the corticotropin-releasing factor system: effects on physiology and behavior.

Genetic modifications of the genes that encode proteins integral to the corticotropin-releasing factor (CRF) system have been employed in the creation of mutant mice that serve as tools for studying the role of this neuropeptide in regulated and dysregulated behaviors and physiology. Overexpression of the CRF peptide and CRF binding protein as well as deletion of the peptide, binding protein, and both known receptors has been achieved and these mouse models have been characterized for anatomical, neuroendocrine, and behavioral sequelae. The profile of results, consistent with current knowledge of CRF function from more traditional assays, indicates that enhancement of CRF function is associated with an activation of the hypothalamic-pituitary-adrenal axis, an anxious phenotype, alterations in cognitive performance and reductions in feeding. In general, blockade of CRF function produces the opposite effects. Genetic mouse models allow further analysis of specific elements in the CRF circuitry for which more traditional tools have not existed. These animal models are valuable for increasing our understanding of the underlying pathology associated with a variety of psychiatric and neuroendocrine disorders and for the development and testing of novel treatment agents.

Dissociation of locomotor activation and suppression of food intake induced by CRF in CRFR1-deficient mice.

Corticotropin-releasing factor (CRF) systems are involved in locomotor and feeding behaviors. Two distinct CRF receptor subtypes, CRFR1 and CRFR2, are thought to mediate CRF actions in the central nervous system. However, the role for each receptor in locomotor activity and feeding remains to be determined. Using CRFR1 null mutant mice, the present study examined the functional significance of this receptor in ambulation and feeding. CRF treatment of wild-type mice resulted in increased levels of locomotion whereas no change was observed in CRFR1-deficient mice as compared to vehicle-treated mutant mice. In contrast, CRF decreased food-water intake in both wild type and CRFR1-deficient mice equally. These results support an important role for CRFR1 in mediating CRF-induced locomotor activation, whereas other receptor subtypes, likely CRFR2, may mediate the appetite-suppressing effects of CRF-like peptides.

Mice deficient for corticotropin-releasing hormone receptor-2 display anxiety-like behaviour and are hypersensitive to stress.

Corticotropin-releasing hormone (Crh) is a critical coordinator of the hypothalamic-pituitary-adrenal (HPA) axis. In response to stress, Crh released from the paraventricular nucleus (PVN) of the hypothalamus activates Crh receptors on anterior pituitary corticotropes, resulting in release of adrenocorticotropic hormone (Acth) into the bloodstream. Acth in turn activates Acth receptors in the adrenal cortex to increase synthesis and release of glucocorticoids. The receptors for Crh, Crhr1 and Crhr2, are found throughout the central nervous system and periphery. Crh has a higher affinity for Crhr1 than for Crhr2, and urocortin (Ucn), a Crh-related peptide, is thought to be the endogenous ligand for Crhr2 because it binds with almost 40-fold higher affinity than does Crh. Crhr1 and Crhr2 share approximately 71% amino acid sequence similarity and are distinct in their localization within the brain and peripheral tissues. We generated mice deficient for Crhr2 to determine the physiological role of this receptor. Crhr2-mutant mice are hypersensitive to stress and display increased anxiety-like behaviour. Mutant mice have normal basal feeding and weight gain, but decreased food intake following food deprivation. Intravenous Ucn produces no effect on mean arterial pressure in the mutant mice.

Genetic differences in response to novelty and spatial memory using a two-trial recognition task in mice.

A two-trial memory task, based on a free-choice exploration paradigm in a Y-maze, was previously developed to study recognition processes in Sprague-Dawley rats. Because this paradigm avoids the use of electric shock or deprivation that may have nonspecific effects and does not require learning of a rule, it may be particularly useful for studying memory in mice. Four inbred strains (Balb/cByJ, DBA/2J, C57BL/6J, and SJL/J), an F1 hybrid (C57BL/6 x SJL/J), and one outbred strain (CD1) were used to validate this task in mice and to characterize a strain distribution in response to novelty and working memory. Exploration was measured with a short (2 min) intertrial interval (ITI) between acquisition and retrieval, while memory was examined with longer intervals (30 min, 1 h, and 2 h). A study of the time course of the response to novelty revealed varying degrees of preference and/or habituation to novelty among the different strains, with CD1 exhibiting a very high response to novelty and others showing lower (C57 x SJL hybrids) to complete absence (SJL) of exploration of novelty. Memory span, assessed with increasing ITIs, varied widely among strains from 30 min (C57 x SJL hybrids) to at least 2 h (C57 and BALB). Such demonstrated sensitivity to a wide range of behavioral phenotypes supports the use of this spatial memory task as an effective tool for the study of genetic influences on the response to novelty and recognition processes in mice.

Abecarnil enhances recovery from diazepam tolerance.

Treatment with diazepam (25 mg/kg; p.o., twice-daily for 17 days) induced tolerance to the anticonvulsant effect of diazepam against bicuculline-induced convulsions in mice. Cross-tolerance was observed to the anticonvulsant action of clonazepam, imidazenil but not abecarnil. While substitution of clonazepam (12 mg/kg; p.o., twice-daily for 15 days) for diazepam did not affect tolerance to diazepam, substitution of imidazenil (17 mg/kg; p.o., twice-daily for 15 days) for diazepam significantly increased sensitivity to the anticonvulsant effect of diazepam, although tolerance was not abolished. Tolerance to diazepam progressively decreased either after suspension of diazepam administration or replacement treatment with abecarnil (20 mg/kg; p.o., twice-daily). Complete recovery of diazepam efficacy was detected after 8 and 15 days of administration of abecarnil and vehicle, respectively. Binding experiments using [3H]-flumazenil showed that Kd values did not differ among treatment groups. A significant decrease in Bmax (-42%) was observed in the cortex of diazepam-tolerant mice whether or not also treated with imidazenil and clonazepam. Conversely, chronically diazepam-treated mice, that further received abecarnil for either 8 or 15 days or vehicle for 15 days showed Bmax values similar to those of vehicle-treated mice never exposed to diazepam. Results suggest that repeated abecarnil administration to diazepam-tolerant mice can facilitate re-adaptation of receptors to the diazepam-free state. It is proposed that replacement therapy with abecarnil after long-term treatment with conventional benzodiazepines (BDZs) may provide a novel approach for reducing tolerance to their anticonvulsant effects.

Reduced anxiety-like and cognitive performance in mice lacking the corticotropin-releasing factor receptor 1.

Corticotropin-releasing factor (CRF) has been hypothesized to be involved in the pathophysiology of anxiety, depression, cognitive and feeding disorders. Two distinct CRF receptor subtypes, CRFR1 and CRFR2, are thought to mediate CRF actions in the CNS. However, the role for each receptor subtype in animal models of neuropsychiatric disorders remains to be determined. Using CRFR1 deficient mice, the present study investigated the functional significance of this CRF receptor subtype in anxiety-like and memory processes. CRFR1 knockout mice displayed an increased exploratory behavior in both the Elevated Plus-maze (EPM) and the Black and White (B-W) test box models of anxiety, indicating an anxiolytic-like effect of the CRFR1 gene deletion. In contrast, during the retrieval trial of a two-trial spatial memory task wild type mice made more visits to and spent more time in the novel arm as opposed to the two familiar ones of a Y-maze apparatus. No increase in the level of exploration of the novel arm by the CRFR1 deficient mice was observed. This indicates that CRFR1 knockout mice are impaired in spatial recognition memory. These results demonstrate that genetic deletion of the CRFR1 receptor can lead to impairments in anxiety-like and cognitive behaviors, supporting a critical role for this receptor in anxiety and cognitive biological processes.

A new place conditioning paradigm to study tolerance to opiates in mice.

Tolerance to the rewarding properties of morphine was investigated in mice using a new conditioned place preference (CPP) procedure. Four pairings of morphine with specific environmental cues induced a significant CPP for the drug-paired cues. Further opiate conditioning trials in the presence of the same environmental cues revealed no change in the drug-induced CPP on repeated test sessions. Subsequent exposure of the same animals to conditioning trials by pairing morphine with a set of novel environmental cues showed that the opiate was still able to produce a CPP in mice treated with a total of 16 morphine injections. The present CPP paradigm may prove useful to investigate tolerance to the rewarding properties of drugs of abuse.

The expression of neuropeptide-induced excessive grooming behavior in dopamine D1 and D2 receptor-deficient mice.

Grooming behavior in rodents has long been related to dopamine receptors in the brain. However, the relative contribution of dopamine D1-like receptors (D1 and D5) and D2-like receptors (D2, D3 and D4) in this behavior has not been established yet. Spontaneous novelty-induced grooming (as assessed with a 30-min sampling test) was reduced in knockout mice lacking the dopamine D1, receptor. Furthermore, the intracerebroventricular (i.c.v.) injection of small quantities of oxytocin, prolactin or the adrenocorticotrophic hormone 1-24 fragment, ACTH-(1-24) was followed by a diminished level of novelty-induced excessive grooming. These neuropeptides caused a sustained increase in grooming level of control animals (wild type). Interestingly, the i.c.v. injection of beta-endorphin enhanced novelty-induced grooming to a level similar in control and knockout mice. The systemic administration of the dopamine D2 receptor antagonist, sulpiride did not suppress the residual grooming activity shown by animals injected with oxytocin, prolactin or ACTH-(1-24), and did not change the behavioral expression of those injected with beta-endorphin. In contrast, the systemic administration of the opioid receptor antagonist, naloxone, totally suppressed the residual grooming activity of oxytocin-, prolactin- or ACTH-(1-24)-injected mice and of those treated with beta-endorphin. In contrast with the behavioral deficit observed in dopamine D1 receptor-deficient mice, dopamine D2 receptor-null animals showed a normal expression of spontaneous novelty-induced grooming and a high level of grooming activity induced by i.c.v. injection of oxytocin, prolactin, ACTH-(1-24) or beta-endorphin. Again, the peripheral injection of naloxone was followed by a suppression of neuropeptide-induced excessive grooming in these animals. These data suggest that dopamine D1 receptors are involved in the expression of novelty-induced grooming in mice. In contrast, dopamine D2 receptors seem not to be important for the expression of this behavior. Furthermore, neuropeptide-enhanced grooming involves dopamine D1, but not dopamine D2 receptors. However, neurotransmitters other than dopamine (e.g., endorphins) may play a supplementary role in neuropeptide-enhanced grooming in mice.

Understanding corticotropin releasing factor neurobiology: contributions from mutant mice.

Mice with transgenic expression or deletion of the CRF peptide, transgenic expression of the CRF-BP or deletion of specific CRF receptor subtypes exist and will be valuable for examining candidate mediators in animal model systems recapitulating a variety of normal function. In particular, results described in this review implicate CRF in acute emotional responses studied in animal models of anxiety and drug abstinence. CRF also appears to play a role in behavioral and physiological plasticity judging by alterations in HPA reactivity to stress, information processing and energy balance regulation in CRF mutant models. Accordingly, the creation of genetically engineered mice now permits the evaluation of contributory roles for several CRF-related gene products in the pathophysiology of a variety of complex behavioral disorders. For example, the postulated causal linkage between overactivation of CRF systems and the hyper-emotionality which characterizes human affective disorders can now be more thoroughly evaluated by examining the phenotype of CRF mutant mice in animal models of depression, dementia and substance abuse.

Corticotropin releasing factor receptor 1-deficient mice display decreased anxiety, impaired stress response, and aberrant neuroendocrine development.

Corticotropin releasing factor (CRF) is a major integrator of adaptive responses to stress. Two biochemically and pharmacologically distinct CRF receptor subtypes (CRFR1 and CRFR2) have been described. We have generated mice null for the CRFR1 gene to elucidate the specific developmental and physiological roles of CRF receptor mediated pathways. Behavioral analyses revealed that mice lacking CRFR1 displayed markedly reduced anxiety. Mutant mice also failed to exhibit the characteristic hormonal response to stress due to a disruption of the hypothalamic-pituitary-adrenal (HPA) axis. Homozygous mutant mice derived from crossing heterozygotes displayed low plasma corticosterone concentrations resulting from a marked agenesis of the zona fasciculata region of the adrenal gland. The offspring from homozygote crosses died within 48 hr after birth due to a pronounced lung dysplasia. The adrenal agenesis in mutant animals was attributed to insufficient adrenocorticotropic hormone (ACTH) production during the neonatal period and was rescued by ACTH replacement. These results suggest that CRFR1 plays an important role both in the development of a functional HPA axis and in mediating behavioral changes associated with anxiety.

Conditioned place preference: no tolerance to the rewarding properties of morphine.

The effect of repeated morphine administration on conditioned place preference (CPP) using a novel treatment schedule, i.e., drug treatment was always contingent with the conditioned environmental stimuli, was investigated. We also examined whether changes in the mu- and kappa-opioid receptor binding occurred in the brain of morphine-treated animals. Intraperitoneal (i.p.) administration of morphine (2 and 10 mg/kg) induced a place preference after 8 daily conditioning trials (4 morphine injections on alternate trials), the level of preference being the same with the two doses of the opiate. No change in place preference was observed in the morphine-treated rats at 2 mg/kg, when animals were further trained up to a total of 32 conditioning trials (16 morphine injections). Conversely, after 20 conditioning trials (10 morphine injections), a stronger CPP response developed in the morphine-treated rats at 10 mg/kg. Signs of morphine withdrawal were never detected in morphine-treated rats during the experiment. Loss of body weight (index of opiate dependence) was not observed either 24 h or 48 h after the last morphine administration. mu- and kappa-opioid receptor density and affinity were not affected by repeated morphine administrations at either dose. The results demonstrate that no tolerance develops to the rewarding properties of morphine. Indeed, a sensitisation effect may occur at increasing doses of the opiate. Furthermore, changes in the rewarding effect of morphine are not dependent upon alterations in opioid receptors involved in the reinforcing mechanisms.

Effects of acute or chronic administration of risperidone on motor and sexual behavior of male rats.

A number of experiments were carried out to explore the behavioral profile of a novel antipsychotic, risperidone, after acute or chronic administration, in a dose range of 0.1-10 mg kg-1. This drug did not affect the acquisition and retention of avoidance behaviors in a dose of 0.1 mg kg-1, either after acute or chronic administration. Higher doses induced a inhibited acquisition and a facilitated extinction (only after chronic treatment) of active avoidance behavior, but no significant effect on the retention of passive avoidance responses. In contrast, haloperidol inhibited the acquisition and facilitated the extinction of active avoidance behavior, and reduced the retention of passive avoidance reaction at the dose of 0.1 mg kg-1 injected either acutely or chronically. Ambulation and rearing of rats rated in an open field was increased by risperidone injected acutely at the dose of 1 mg kg-1. Under the same experimental conditions, grooming appeared to be reduced. In the same test, acute or chronic haloperidol 1 or 10 mg kg-1 inhibited all behavioral items. Furthermore, in contrast to haloperidol, the acute or chronic administration of risperidone in a dose range of 0.1-10 mg kg-1 did not substantially induce catalepsy and did not affect apomorphine-induced stereotypies. Also, the dose of 0.1 mg kg-1 induced a facilitation of male sexual behavior by increasing the frequency and reducing the latency of mountings, intromissions and ejaculations, while haloperidol 1 or 10 mg kg-1 inhibited this behavior. These findings suggest that the pharmacological profile of risperidone differs from that of classical neuroleptics, like haloperidol, probably due to different mechanism or site of action.

Abecarnil, a beta-carboline derivative, does not exhibit anticonvulsant tolerance or withdrawal effects in mice.

Development of tolerance and dependence has been reported to occur upon chronic administration of traditional benzodiazepines (BZDs). We compared the effect of chronic treatment with abecarnil, a beta-carboline derivative with high affinity for central BDZ receptors, and diazepam, the BDZ prototype, in mice. After acute administration, abecarnil was as potent and effective as diazepam in protecting from bicuculline-induced convulsion. The time-course analysis of two peak equieffective doses of abecarnil (1.9 mg/kg p.o.) and diazepam (2.7 mg/kg p.o.) showed a similar duration of action. The anticonvulsant potency of diazepam was reduced in mice given chronic diazepam (25 mg/kg p.o., 2 times a day for 17 days). No tolerance to abecarnil was apparent when the drug was administered for the same period using a comparable dose (20 mg/kg p.o.). Severe symptoms of precipitated withdrawal were observed upon administration of the BDZ partial inverse agonist Ro 15-3505 in mice treated chronically with diazepam but not abecarnil. In mice made tolerant to diazepam, maximum [3H]-flumazenil binding sites were reduced in both cerebral cortex (-50%) and cerebellum (-55.2%). No changes in [3H]-flumazenil binding were measured in chronic abecarnil-treated mice. These data indicate that abecarnil possesses a very low tolerance/dependence liability and does not affect BZD receptor density after chronic administration.

Lack of anticonvulsant tolerance and benzodiazepine receptor down regulation with imidazenil in rats.

1. Development of anticonvulsant tolerance and benzodiazepine (BZD) receptor down-regulation has been reported to occur upon chronic administration of conventional BZDs. We compared the effect of chronic treatment with imidazenil, a new BZD partial agonist, and diazepam in rats. 2. After acute administration, imidazenil was more potent though less effective than diazepam in protecting from bicuculline-induced seizure. The time-course analysis of two peak equieffective doses of imidazenil (2.5 mumol kg-1 p.o.) and diazepam (35 mumol kg-1, p.o.) showed a longer lasting action of the former drug. 3. The anticonvulsant efficacy of diazepam (35 mumol kg-1, p.o.) was reduced in rats given chronic diazepam (35 mumol kg-1 p.o., 3 times a day for 8-15 days). No tolerance to imidazenil (2.5 mumol kg-1, p.o.) was apparent after 130-day administration with imidazenil (2.5 mumol kg-1, p.o., 3 times a day). 4. Plasma levels of imidazenil and diazepam, assessed 30 min after administration, were not changed in chronically treated animals. 5. In rats made tolerant to diazepam, the maximum number of [3H]-flumazenil binding sites were reduced in both cerebral cortex (-36%) and cerebellum (-42%). No changes in [3H]-flumazenil binding were found in chronic imidazenil-treated rats. 6. Specific [3H]-flumazenil binding in vivo was decreased in the forebrain of chronic diazepam- but not of chronic imidazenil-treated animals. 7. These data indicate that imidazenil possesses a very low tolerance potential to its anticonvulsant activity and does not affect BZD receptor density even after prolonged administration.

[Description of a case of pyogenic spondylodiscitis with bacterial metastatic colonization]. / Descrizione di un caso di spondilodiscite piogenica con colonizzazioni batteriche metastatiche.

A case of staphylococcus spondylodiscitis with vertebral collapse and recurrent bacterial colonizations at pulmonary and gluteal level consequent on a septicaemic condition that arose as a result of a missed diagnosis is reported. The misleading factor was the pre-existing spondylarthrosis and marked osteoporosis.