The cerebellum monitors errors and entrains executive networks.

Frontal midline θ (Fmθ) activity occurs in medial prefrontal cortices (mPFC), when expected and actual outcomes conflict. Cerebellar forward models could inform the mPFC about this mismatch. To verify this hypothesis we correlated the mPFC activation during a visuomotor tracking task (VM) with performance accuracy, in control and cerebellum-lesioned participants. Additionally, purely visual (V), motor (M) and a motor plus visual tasks (V + M) were performed. An Independent Component, with a mid-frontal topography scalp map and equivalent dipole location in the dorsal anterior cingulate cortex accounted for Fmθ. In control participants Fmθ power increased during VM, when the error level crossed a threshold, but not during V + M, M and V. This increase scaled with tracking error. Fmθ power failed to increase during VM in cerebellar participants, even at highest tracking errors. Thus, in control participants, activation of mPFC is induced when a continuous monitoring effort for online error detection is required. The presence of a threshold error for enhancing Fmθ, suggests the switch from an automatic to an executive tracking control, which recruits the mPFC. Given that the cerebellum stores forward models, the absence of Fmθ increases during tracking errors in cerebellar participants indicates that cerebellum is necessary for supplying the mPFC with prediction error-related information. This occurs when automatic control falters, and a deliberate correction mechanism needs to be triggered. Further studies are needed to verify if this alerting function also occurs in the context of the other cognitive and non-cognitive functions in which the cerebellum is involved.

Intramuscular injections in newborns: analgesic treatment and sex-linked response.

AIM: To compare the analgesic effect of three treatments to relieve the pain produced by intramuscular injections (IMI) in term newborns, and to assess sex-linked differences in their response to pain. MATERIAL AND METHODS: We studied 62 babies. Each baby received antibiotic IMIs for clinical aims. During each IMI, one of the following analgesic treatments was utilized: oral 33% glucose (OG), sensorial saturation (SS), or topic anesthetic cream (TAC). SS is a validated analgesic method, based on the combination of three stimulations (tactile, acoustic and gustative). During the IMI, pain level was assessed with the use of the DAN scale, a validated neonatal pain scale. All babies who received three distinct analgesic procedures for three distinct IMIs were enrolled. Mean pain scores of the three analgesic treatment groups were compared. We then compared mean pain scores of females vs males in the whole cohort and within each treatment group. RESULTS: The 95% Confidence Intervals of pain scores were 5.6-6.5 for TAC, 1.4-2.3 for OG and 0.6-1.2 for SS: when treated with TAC, babies' pain scores were significantly higher than with OG or SS (p <0.0001); when treated with OG, babies' pain scores were higher than SS (p = 0.002). Females' mean pain score was significantly higher than males' mean pain score: (95% CI: 2.9-4.1 vs 2.0-3.1; p = 0.01). OG and SS produced significantly higher mean DAN scores in females than in males. Also in the TAC group females' mean DAN scores were higher than males, though this last difference was not statistically significant. CONCLUSION: This is the first study to show the effectiveness of nonpharmacologic analgesia in relieving IMI pain. It is also the first study to clearly show that the sex differences in pain perception are present since birth.

Synthesis of esters of androgens with unsaturated fatty acids for androgen requiring therapy.

Androgens' metabolism and activity are gaining a more and more important role in human physiology particularly referring to aging and to neurodegenerative diseases. Androgen treatment is often required for long-lasting disorders. In order to improve their duration and effects, androgens can be administered as esters of carboxylic acids. The novelty of our research is the use of esters of androgens with specific unsaturated fatty acids, in order to reduce possible side effects particularly related to chronic pathologies with altered lipid homeostasis such as X-linked adrenoleukodystrophy and cardiovascular disorders. Thus the esters of the main androgenic substances testosterone, dihydrotestosterone (DHT) and their metabolite 5α-androstan-3α,17ß-diol were chemically obtained by coupling with different unsaturated fatty acids. To this aim, fatty acids with various degree of unsaturation and belonging to different series were selected. Specifically, oleic acid (18:1, n-9), linoleic acid (18:2, n-6), and the n-3 fatty acids, α-linolenic acid (18:3), eicosapentaenoic acid (EPA, 20:5), and docosahexaenoic acid (DHA, 22:6) were used obtaining corresponding esters with acceptable yields and good degree of purity. All the synthesized compounds were tested for their cytotoxic activities in mouse NIH3T3 and human astrocyte cell lines. The esters demonstrated good tolerability and no in vitro cytotoxic effect in both cell cultures. After these promising preliminary results, the esters will be suitable for in vivo studies in order to ascertain their pharmacokinetic characteristics and their biological effects.

The soy phytoestrogens genistein and daidzein as neuroprotective agents against anoxia-glucopenia and reperfusion damage in rat urinary bladder.

Some bladder disorders, such as obstructive bladder and hyperactivity, may be caused partly by ischemia/reperfusion injury (I/R). The neuroprotective effects of estrogens were demonstrated in in vitro studies and a great interest in soy isoflavones (genistein and daidzein) as alternative to the synthetic estrogen receptor modulators for therapeutic use has been pointed out. The aim of this study was to investigate the effect of genistein and daidzein, on rat detrusor smooth muscle contractility and their possible neuroprotective role against I/R-like condition. Whole rat urinary bladders were subjected to in vitro anoxia-glucopenia (A-G) and reperfusion (R) in the absence or presence of drugs and response to electrical field stimulation (EFS) of intrinsic nerves evaluated. Furthermore rats were treated in vivo for 1 week with the phytoestrogens and the same in vitro protocol was applied to the ex vivo bladders. Antioxidant activity of genistein and daidzein on the A-G/R model was determined by measuring malonyldialdehyde (MDA). Moreover, hormones plasma levels were determined by radioimmunoassay. Genistein and daidzein administered either in vitro or in vivo showed significant neuroprotective effect and antioxidant activity. Testosterone and 17ß-estradiol plasma levels were not modified by daidzein, while a significant decrease of testosterone in genistein treated rats was evident. Moreover both phytoestrogens significantly decreased detrusor contractions induced by EFS in a concentration-dependent manner. For being either neuroprotective and myorelaxant, genistein and daidzein could be considered a good lead for new therapeutic agents to protect the urinary bladder from hyperactivity and nerve damage.

Can hyperbaric oxygenation therapy (HOT) modify the blood testosterone concentration?

INTRODUCTION: Testosterone has a modulating effect on inflammatory and healing processes. In this study, we evaluate whether hyperbaric oxygenation therapy (HOT) modifies the blood concentration of total testosterone (TT) in patients treated for different pathologies. MATERIALS AND METHODS: Fourteen male patients (23-72 years old) were treated with 90-min HOT sessions (range 4 to 23 sessions) as an adjuvant to the following conditions: leg fractures, osteonecrosis, diabetic foot, firearm injuries, complicated arthroprosthesis and underwater diving embolism. As controls, six healthy male volunteers (37-51 years old) were subjected to 10 HOT sessions. Testosterone plasma levels were determined immediately before the first HOT session and the day after the last session. RESULTS: At the end of treatment, 12 patients fully recovered and 2 (diabetic foot patients) showed a marked improvement. Testosterone significantly increased after hyperbaric oxygenation therapy in both patients and controls (ANOVA, p<0.004). DISCUSSION: We conclude that hyperbaric oxygenation therapy increases the blood concentration of total testosterone in patients as well as in healthy men. This finding raises new questions and indicates the need to investigate the causes of this increase and its therapeutic significance. Since testosterone modulates inflammation and healing processes, it is possible that hormonal changes are the mechanisms affected by hyperbaric oxygenation therapy.

Glucocorticoid receptor mRNA expression in peripheral blood mononuclear cells in high trained compared to low trained athletes and untrained subjects.

BACKGROUND: Physiological needs during prolonged exercise are a potent stimulus for the hypothalamic-pituitary-adrenal (HPA) axis. Hence, athletes undergoing daily endurance training sessions may have frequent and prolonged phases of endogenous hypercortisolism. Since chronic glucocorticoids treatment leads to down-regulation of glucocorticoid receptor alpha (GR-alpha) mRNA expression, endurance training could lead to modulation of GR expression. AIM: The aim of the study was to evaluate GR-alpha and GR-beta mRNA expressions in peripheral blood mononuclear cells and plasma cortisol, ACTH and cortisol binding globulin (CBG) concentrations at rest in subjects undergoing different training regimes. SUBJECTS AND METHODS: Nine high trained (HT) swimmers (training volume: 21.6+/-1.7 hours/week in 10-12 sessions) were compared with two age-matched control groups represented by 8 low trained (LT) runners (training volume: 6.4+/-2.6 h/week in 3-5 sessions) and 9 untrained subjects. Expression of GR was determined by RT-PCR of total RNA. Hormone levels were determined by radioimmunoassay methods. RESULTS: HT athletes showed 10 times less GR-alpha mRNA expression than the untrained subjects, while LT athletes exhibited values about twofold less than the untrained subjects. GR-beta mRNA expression was undetectable in all subjects. No differences were observed among the three groups in hormone levels. CONCLUSIONS: GR- alpha mRNA expression is repressed in proportion to the amount and frequency of the stressful stimuli due to training. Hence, this down-regulation may be a consequence of the frequent and prolonged exposure to cortisol acute elevations induced by training. GR-beta did not play an important role in inducing the down-regulation of GR-alpha mRNA expression observed.

[Sexual rehabilitation with intracavernous PGE1 injections and oral drug administration in diabetic patients non-responder to oral therapy alone].

Diabetes is an important risk factor in erectile dysfunction (ED), acting via several mechanisms. We assessed the efficacy of intracavernous injections (ICI) rehabilitation and oral systematic therapy in diabetic patients, as well as the response of controls to oral therapy 'on demand'. MATERIALS AND METHODS. Sixteen diabetic patients with ED were treated with vasoactive drugs orally when needed, without satisfactory erections. The patients underwent then ICI rehabilitation with PGE1 20 mcg twice weekly for 4 weeks, followed by the administration of oral drugs twice weekly for 4 weeks. Before and after rehabilitation, the patients completed a detailed anamnestic protocol to study their libido (always present); they answered questions Q3 and Q4 of the IIEF questionnaire. During ICI, a study with dynamic echocolordoppler (ECCD) was carried out. All patients had Type 2 diabetes: 10 were treated with oral antidiabetics, 4 were treated with insulin, and in the other 2 patients, treated with insulin, a sensitive neuropathy of the lower limbs was diagnosed. Fourteen patients were treated with antihypertensive drugs. RESULTS. Before rehabilitation, the mean responses to questions 3 and 4 of the IIEF (International Index of Erectile Function) questionnaire were 1.6 and 1.5 respectively; after rehabilitation, the mean responses were 2.68 and 2.5, respectively. The ECCD test showed an arterial component in 4 cases and a high end-diastolic velocity (EDV) in 14 cases. Four patients (25%), 2 of which had neuropathy, and 2 were in advanced age, did not respond to PGE1 or to oral therapy, 4 patients (25%) (2 treated with insulin and 2 by oral therapy) responded to ICI but not to oral therapy, while 8 patients (50%) showed a good response to both injectable and oral therapy, with good Q3 and Q4 scores. CONCLUSIONS. Good endothelial function appears to be essential for the maintenance of acceptable erectile function. Diabetes has a negative effect on this function, as does hypoxia and low perfusion. Based on the principle that a good erection improves endothelial function, we tried to determine if oral systematic and intracavernous rehabilitation would improve erectile function in diabetic patients. The results indicate that diabetes interferes with erectile function, compromising the effects of the vasoactive drugs. However, integrated systematic rehabilitation appears to allow a good erectile response to both intracavernous and oral therapy in a large number of cases. Therefore, we support this kind of rehabilitative protocol in the treatment of ED in diabetic patients.

Single opioid administration modifies gonadal steroids in both the CNS and plasma of male rats.

While morphine remains one of the most widely used opioids for the treatment of painful conditions, other opioids are also commonly employed. Because of the interactions between opioids and gonadal hormones, in particular opioid-induced hypogonadism, this study investigated the effects of widely used opioids on plasma testosterone and estradiol levels and brain testosterone levels in male rats. Animals were s.c. injected with two concentrations of morphine (5 or 10 mg/kg), fentanyl (0.05 or 0.1 mg/kg), tramadol (10 or 40 mg/kg), buprenorphine (0.05 or 0.1 mg/kg) or saline (0.7 ml/kg). Four or 24 h after treatment, the rats were deeply anesthetized to collect blood samples from the abdominal aorta and to perfuse the brains with saline. Plasma and brain hormone levels were measured by radioimmunoassay. In rats studied 4 h after treatment, all the opioids except tramadol 10 mg/kg decreased plasma testosterone in comparison with saline administration. At the same time, plasma estradiol levels were lower than control in the groups treated with the low doses of morphine, tramadol and buprenorphine, while estradiol remained at control levels in the other groups. Twenty-four hours after treatment, plasma testosterone levels were different (higher) than control in the animals treated with the low doses of morphine, fentanyl and buprenorphine. Estradiol was lower than control in the low dose groups, while the high doses did not produce any changes with respect to control. Four hours after treatment, brain testosterone was drastically decreased in all groups except buprenorphine, in which it remained at control levels. All groups returned to control levels at 24 h after treatment. In conclusion, opioids exert important effects on plasma and CNS sex hormone levels. The different magnitude and time-course of the effects of the different opiates on testosterone and estradiol levels are likely due to their different mechanism of action.

Effects of gonadectomy and pain on interferon-gamma production in splenocytes of male and female rats.

The role of gonadal hormones and persistent pain (formalin test) in the regulation of interferon-gamma (IFN-gamma) production in splenocytes was investigated in male and female rats. Animals were either sham-operated (Intact) or gonadectomized (GDX) and, 3 weeks later, were subcutaneously injected with formalin (50 microl, 10%) or only pricked with a syringe needle in the dorsal hind paw. Sixty minutes after treatment the animals were deeply anesthetized and the spleens were dissected under aseptic conditions. Blood was collected from the abdominal aorta for measurement of plasma steroids. IFN-gamma production was determined in vitro in the splenocytes after Con A stimulation. Splenocytes of Intact females showed higher IFN-gamma production than those of Intact males. This sex difference disappeared in GDX animals because of the lower levels in GDX females. Formalin decreased IFN-gamma in both Intact and GDX groups. In females, there was a positive correlation between IFN-gamma production in splenocytes and plasma estradiol levels. The present data demonstrate a sex difference in IFN-gamma production (due to the immunostimulating effect of estradiol in females) and an immunodepressive role of pain in both sexes.

Bisphenol-A differently affects estrogen receptors-alpha in estrous-cycling and lactating female rats.

The effect of long-term exposure to bisphenol-A (BPA) on estrogen receptor-alpha (ER) immunoreactivity was studied in the medial preoptic area, arcuate nucleus and the ventromedial nucleus of the hypothalamus of estrous cycling and lactating female rats. Pregnant/lactating or estrous cycling rats were exposed to BPA (40 mg/Kg/day) or peanut oil. Lactating females showed fewer ER-immunoreactive cells than non-lactating females in the medial preoptic area and ventromedial nucleus of the hypothalamus. BPA induced an increase in ER-immunoreactive cells in the medial preoptic nucleus irrespective of lactation. BPA only induced a decrease in ER-immunoreactive cells in the arcuate nucleus of the lactating group; oil induced an increase in ER-immunoreactive cells in the lactating with respect to non-lactating group. The results demonstrate that exposure of adult females to BPA modifies the number of ERs.

Effects of gonadal hormones and persistent pain on non-spatial working memory in male and female rats.

There are indications of a modulatory role carried out by gonadal hormones and pain in cognitive functions. We have examined this issue in male and female rats by assessing the impact of gonadectomy and persistent pain on the object recognition test. Intact and gonadectomized male and female rats were exposed to an open field (15 min) in which three objects were placed (Trial 1); the same test was repeated 2 h later (Trial 2), after the replacement of a "familiar" object with a novel one. Three days later (Day 2), the same procedure was repeated (Trial 3 and 4 with 2 h in between) but half of the animals were exposed to formalin-injection immediately before Trial 3. The latency, frequency and duration of approaching the three objects were recorded in each trial and compared by sex, gonadectomy and formalin treatment. The results showed that gonadectomized males and females had lower levels of approach to all objects and less locomotor/exploratory activity than intact animals in all experimental trials; their behaviour was not affected by repetition of the test or by pain. On Day 1, intact males showed a higher level of approach to the novel object than females. In intact males, the 2 h delay between the first and second trial failed to induce any significant modification of exploration of the novel object with respect to the familiar one, while in intact females the novel object was approached much less than the familiar one. Similarly on Day 2, the novel object was approached for a longer time by intact males than by all the other groups. In conclusion, our data show that physiological levels of circulating gonadal hormones significantly affected the performance of male but not female rats when exposed to the object recognition test.

Role of gonadal hormones in formalin-induced pain responses of male rats: modulation by estradiol and naloxone administration.

The aim of this study was to assess the possible mediation of endogenous opioids in the effects of gonadal hormones on the responses to formalin pain. We studied the effects of intracerebroventricular injection of estradiol and/or naloxone on the magnitude and time-course of the formalin-evoked behavioural and hormonal responses of intact and gonadectomized male rats. Animals were gonadectomized or left intact; on days 20 and 21 after surgery, they were intracerebroventricularly injected with 17beta-estradiol (1 microg/5 microl) or saline. On day 22, the animals received naloxone (2.5 microg/5 microl) or saline intracerebroventricularly and then, 15 min later, were subcutaneously injected with formalin (50 microl, 5%) or only pricked with a syringe needle in the dorsal hindpaw. The rats were then introduced to a testing apparatus where the formalin-induced licking, flexing and jerking of the injected limb and the other spontaneous behaviours were recorded for 60 min. At the end of the test, the animals were killed and blood was collected from the trunk. Gonadectomy and naloxone increased flexing duration independently of the other treatments. In gonadectomized rats, estrogen increased licking duration and decreased paw-jerk frequency during the first phase (0-15 min) of the formalin test. During the second phase (16-60 min), licking was increased by estrogen only in intact animals. Treatment with naloxone completely abolished all these modifications. The three measures of activity (rearing, inner and outer crossing) showed that while in sham-treated animals the gonadectomy-induced decrease in activity was completely counteracted by estrogen administration, in formalin-treated animals the gonadectomy-induced decrease was not affected by estrogen. In fact, estrogen appeared to further depress the motor activities in the formalin groups. Naloxone reversed these modifications only for outer crossing frequency, blocking the gonadectomy-induced decrease in sham-treated animals. Corticosterone plasma levels were increased by formalin only in estrogen-treated animals, independently of naloxone. In conclusion, these data indicate an important role of both male gonadal hormones and estrogen in formalin-pain responses, acting through opiate and non-opiate mechanisms.

Gonadectomy and persistent pain differently affect hippocampal c-Fos expression in male and female rats.

Hippocampal c-Fos expression was studied in male and female rats after gonadectomy and persistent pain. Three weeks after surgery, animals were sham- or formalin-injected (50 microl, 10%) and placed in a familiar testing apparatus. The formalin-evoked licking, flexing and jerking of the injected paw were recorded for 60 min, c-Fos was determined in the dorsal and ventral hippocampus: dentate gyrus (DG), CA1 and CA3. Gonadectomy induced higher c-Fos in the dorsal DG of both sexes, in all ventral subfields of males and in the ventral CA3 of females. In normal males and females, formalin increased c-Fos in the dorsal DG and in the male ventral subfields. In gonadectomized ones formalin decreased or did not change c-Fos. Gonadectomy induced longer flexing in males and females. These data indicate an important and sex-dependent interaction between gonadal hormones, nociceptive input and neuronal activity in the hippocampus.

Effects of formalin pain on hippocampal c-Fos expression in male and female rats.

Immediate early genes are crucial intermediates in a cascade linking membrane stimulation to long-term alterations of neuronal activity. In the present experiment, we performed immunohistochemistry for c-Fos to determine the effects of persistent pain on cells of the hippocampus of male and female rats. Animals were subcutaneously injected with formalin (50 microl, 10%) and perfused: 2 h later, time 2; 24 h later, time 24; 24 h later after 20 min of the open-field test, time 24/OF. Controls were left undisturbed. In control, c-Fos was higher in females than in males in all hippocampal fields. In males at time 2, formalin increased c-Fos in the dentate gyrus (DG) and CA3 fields; at time 24, c-Fos returned to the control level; at time 24/OF, c-Fos was higher than in control in the DG, but not in the other fields. In the formalin-treated females at time 2 and at time 24, c-Fos levels were lower, or tended to be lower, than in control in all hippocampal fields; at time 24/OF, c-Fos levels in the DG were higher than in control and in males. In conclusion, persistent pain had different effects on c-Fos in the hippocampal subfields, depending on the time after treatment and the sex of the subject.

The role of Fbg in platelet adhesion to polymeric materials in conditions of psychological stress.

The effect of psychological stress on platelet adhesion to five polymeric materials (polyurethane, polyurethane filled with BaSO4, polyethyleneterephthalate, silicone and low-density polyethylene) was studied. The platelets were obtained from non-stressed and stressed rabbits as platelet-rich plasma (PRP) and, once washed (Pw), were suspended in different media, i.e. in platelet poor plasma (Pw-PPP), in serum (Pw-S) and in Krebs-Ringer solution (Pw-KR). Scanning electron microscopy of platelet adhesion and morphology revealed differences in the platelet activating power of the various materials. The washing procedure and resuspension in PPP generally resulted in an increased number of adherent platelets, compared with the number of platelets adherent to the same material in PRP. However, platelets washed and suspended in Pw-KR or Pw-S showed the same shape distribution as in PRP. When platelets from stressed rabbits were used, there was very strong aggregation and activation of the platelets in both PRP and Pw-PPP, independent of the chemical nature and surface structure of the material. In contrast, in Pw-KR and Pw-S (in which Fbg is absent) a general picture of single, not very modified platelets was observed. Their number and shapes changed according to the nature of the different materials. On the whole, the present results confirm our original hypothesis of a key role of the psychological condition of the blood donor and strongly indicate Fbg as the determinant factor in the pattern of platelet adhesion.

Emotional behavior in female rabbits: hippocampal EEG and neuroendocrine aspects.

Behavior, hippocampal electrical activity, plasma hormones and hippocampal choline acetyltransferase (ChAT) were considered in two groups of female rabbits, different in age and condition of breeding, both in the presence and absence of emotional stimuli. In the two groups of female rabbits (4 and 18 months old), permanent electrodes were implanted bilaterally in the two dorsal hippocampi and, one week later, the animals were exposed for three consecutive days to the following tests: Day 1, novel environment (NE); Day 2, object with odor (O); Day 3, sparrow hawk (SP). Behavior: in comparison with Day 1, exploration was decreased by the object and by the sparrow hawk in the older females and increased in the younger ones. Quiet and alert immobility was higher during the SP test, while freezing and pointing were lower in the older females than in the young ones. Electrical activity: peak frequency was lower in younger females than in the older ones. The frequency band distribution corresponding to exploration and immobility showed that in the older females the % of high frequency band increased from the first to the third day of testing, while an opposite trend was present in the younger females. In the case of freezing the hippocampal electric activity showed a more rhythmical component in younger females than in the older ones. Hormones: increased corticosterone levels after the SP test were directly correlated with exploration and inversely correlated with freezing. Dorsal hippocampal ChAT was directly correlated with quiet immobility. These data indicate a relationship between the responses to the anxious/fearful stimuli and the age and/or breeding experience of the female rabbits; this is shown by both the hippocampal electrical activity and the behavioral differences between the two groups.

Neuroendocrine and behavioral effects of CRH blockade and stress in male rats.

Our previous data have shown that restraint (RT), a mild nonpainful stressor, acutely impairs nonsocial and social behavior in male rats. Corticotropin-releasing hormone (CRH) is a regulator of these behavioral responses. To evaluate whether CRH mediates the neuroendocrine and behavioral alterations present 24 h after restraint stress, we administered the CRH antagonist alpha-helical CRH(9-41) (alpha-hCRH) intracerebroventricularly to male rats and we compared its effects with those of saline. Twenty-four hours after treatment, nonsocial behaviors were significantly decreased by alpha-hCRH, this effect being independent of RT. Among social behaviors, only introductory activity showed significant differences as a result of both RT and alpha-hCRH. The concentrations of ACTH in the plasma and those of beta-endorphin in the anterior and neurointermediate lobes of the pituitary were affected by alpha-hCRH treatment. The effect on ACTH was simply related to the administration of the alpha-hCRH, while for beta-endorphin, significant interactions between alpha-hCRH and RT were found. On the whole, these results point to the role played by CRH in the control of neuronal mechanisms involved in the stress-induced effects.

The role of plasma proteins and stress in the assessment of hemocompatibility.

The physiological and psychological conditions of subjects supplying blood for hemocompatibility tests significantly affect the behavior of platelets in terms of both adhesion and activation. The responses of platelets to a standard biomaterial, polyethylene (PE), were examined with blood collected from male rabbits both in basal conditions and after stress. Different media were utilized. First, platelet-rich plasma (PRP) was used to obtain a PE response to contact with platelets. Then platelets drawn from PRP were isolated and washed with Krebs-Ringer solution. One aliquot was suspended in serum (Pw-S) where fibrinogen was absent, another aliquot in Krebs-Ringer solution (Pw-KR) (in order to avoid the influence of the plasma proteins on platelets), and a third aliquot in the original plasma from which the platelets were drawn (Pw-PPP) (in order to restore the initial condition of the plasma but with washed platelets). The analysis of platelet adhesion and morphology was performed by Scanning Electron Microscopy (SEM). Differences in platelet adhesion and morphology were observed with four different media in nonstressed animals, with Pw-PPP showing a higher number and Pw-S and PW-KR lower numbers. Platelet morphology indicated low levels of activation. The platelets drawn from stressed subjects could not be counted in either PRP or PPP medium because they were fully aggregated and adhered; in contrast, in Pw-KR and Pw-S, no significant differences were found with respect to nonstressed conditions, and there was little difference in platelet morphology. All of these factors underline the role of plasma proteins, in particular fibrinogen, in the stress-induced activation of platelets.

Oxotremorine-induced modifications of the behavioral and neuroendocrine responses to formalin pain in male rats.

In the present investigation, the antinociceptive effects of the muscarinic cholinergic agonist, oxotremorine, were evaluated in rats using the formalin test. In Expt. 1, two oxotremorine concentrations (0.1 and 0.2 mg/kg) and two administration times (15 and 1 min before formalin injection) were chosen. All spontaneous and formalin-evoked behavioral responses were considered. In Expt. 2, only the higher concentration of oxotremorine (0.2 mg/kg) was administered 15 or 1 min before the formalin test. The animals were killed 15, 30 or 60 min after formalin treatment. Blood was collected from the trunk to determine corticosterone plasma levels. Some brain areas (hypothalamus, septum and periaqueductal gray matter) were dissected for determination of the beta-endorphin content. Oxotremorine induced a dose- and time-dependent reduction of all formalin-evoked responses: licking was decreased during both the first and second phases of the formalin test, flexing was decreased during the second phase by the higher concentration only and paw-jerk was decreased during the first phase by both concentrations. Rearing and line-crossing were significantly decreased by oxotremorine while exploratory activity was only partially reduced; self-grooming was increased. These effects on exploratory activity and self-grooming were abolished by formalin treatment. beta-endorphin content in the septum was increased by oxotremorine administered 15 min, but not 1 min, before formalin-treatment. beta-endorphin in the hypothalamus increased in all formalin-treated groups independently of oxotremorine administration. These results confirm, and extend to tonic pain, the analgesic effect exerted by oxotremorine on phasic responses. Because of the different effects on each formalin-induced response, they also indicate both spinal and supraspinal CNS sites of action.