The interaction of meal-related, rhythmic and homeostatic mechanisms and the generation of thirst and drinking

One of the primary goals of the study of thirst is to understand why drinking occurs under ad libitum or natural conditions. An appreciation of the experimental strategies applied by physiologists studying thirst from different perspectives can facilitate progress toward understanding the natural history of drinking behavior. Drinking research carried out using three separate perspectives - homeostatic, circadian rhythms, and food-associated - generates types of information about the mechanisms underlying drinking behavior. By combining research strategies and methods derived from each of these approaches, it has been possible to gain new information that increases our appreciation of the interactions between homeostatic mechanisms and circadian rhythms in the modulation of water intake and how these might be related to drinking associated with food intake under near natural conditions

Proliferation of differentiated glial cells in the brain stem

Classical studies of macroglial proliferation in muride rodents have provided conflicting evidence concerning the proliferating capabilities of oligodendrocytes and microglia. Furthermore, little information has been obtained in other mammalian orders and very little is known about glial cell proliferation and differentiation in the subclass Metatheria although valuable knowledge may be obtained from the protracted period of central nervous system maturation in these forms. Thus, we have studied the proliferative capacity of phenotypically identified brain stem oligodendrocytes by tritiated thymidine radioautography and have compared it with known features of oligodendroglial differentiation as well as with proliferation of microglia in the opossum Didelphis marsupialis. We have detected a previously undescribed ephemeral, regionally heterogeneous proliferation of oligodendrocytes expressing the actin-binding, ensheathment-related protein 2'3'-cyclic nucleotide 3'-phosphodiesterase (CNPase), that is not necessarily related to the known regional and temporal heterogeneity of expression of CNPase in cell bodies. On the other hand, proliferation of microglia tagged by the binding of Griffonia simplicifolia B4 isolectin, which recognizes an alpha-D-galactosyl-bearing glycoprotein of the plasma membrane of macrophages/microglia, is known to be long lasting, showing no regional heterogeneity and being found amongst both ameboid and differentiated ramified cells, although at different rates. The functional significance of the proliferative behavior of these differentiated cells is unknown but may provide a low-grade cell renewal in the normal brain and may be augmented under pathological conditions.

The study of genetic polymorphisms related to serotonin in Alzheimer's disease: a new perspective in a heterogenic disorder

Genetic and environmental factors have been implicated in the development of Alzheimer's disease (AD), the most common form of dementia in the elderly. Mutations in 3 genes mapped on chromosomes 21, 14 and 1 are related to the rare early onset forms of AD while the e4 allele of the apolipoprotein E (APOE) gene (on chromosome 19) is the major susceptibility locus for the most common late onset AD (LOAD). Serotonin (5-hydroxytryptamine or 5-HT) is a key neurotransmitter implicated in the control of mood, sleep, appetite and a variety of traits and behaviors. Recently, a polymorphism in the transcriptional control region upstream of the 5-HT transporter (5-HTT) gene has been studied in several psychiatric diseases and personality traits. It has been demonstrated that the short variant(s) of this 5-HTT gene-linked polymorphic region (5-HTTLPR) is associated with a different transcriptional efficiency of the 5-HTT gene promoter resulting in decreased 5-HTT expression and 5-HT uptake in lymphocytes. An increased frequency of this 5-HTTLPR short variant polymorphism in LOAD was recently reported. In addition, another common polymorphic variation in the 5-HT2A and 5-HT2C serotonin receptor genes previously analyzed in schizophrenic patients was associated with auditory and visual hallucinations in AD. These observations suggest that the involvement of the serotonin pathway might provide an explanation for some aspects of the affective symptoms commonly observed in AD patients. In summary, research on genetic polymorphisms related to AD and involved in receptors, transporter proteins and the enzymatic machinery of serotonin might enhance our understanding of this devastating neurodegenerative disorder.

Sleep patterns and sleep-related complaints of Brazilian interstate bus drivers

Sleep-related complaints have become a highlight for physicians as well as public health administrators. Studies of sleep patterns and sleep-related complaints of shift workers have been useful in minimizing reduction in the quality of life due to the warping of the sleep-wake cycle. The objective of the present study was to assess patterns of sleep, sleep-related complaints as well as physical activity and scoring rates for depression and anxiety in interstate bus drivers. Data were obtained with a sleep questionnaire, with the Beck inventory for depression, and the State-Trait Anxiety Inventory (STAI). A total of 400 interstate bus drivers from the northern, southern, central-western and south-eastern regions of Brazil were interviewed. Sixty percent of the subjects interviewed presented at least one sleep-related complaint, 16% admitted to have dozed at the wheel while on duty, and 41% stated that they exercised on a regular basis. Other sleep disturbance complaints reported were: sleep latency 29'17"; physical fatigue, 59.8%; mental fatigue, 45.4%; sleepiness, 25.8%; irritability, 20.6%; insomnia, 37.5%, respiratory disturbances, 19.25% and snoring, 20.75%. Scores for anxiety and depression were not in the pathological range. The present data reinforce the view that bus drivers are generally discontent with shift work and its effects on sleep. Consequently, it is very important to establish an appropriate work schedule for drivers, besides implementing photo-therapy and physical activities in order to minimize sleepiness when driving.

Abnormal proliferative response of the carotid artery of spontaneously hypertensive rats after angioplasty may be related to the depolarized state of its smooth muscle cells

Hypertension is one of the major precursors of atherosclerotic vascular disease, and vascular smooth muscle abnormal cell replication is a key feature of plaque formation. The present study was conducted to examine the relationship between hypertension and smooth muscle cell proliferation after balloon injury and to correlate neointima formation with resting membrane potential of uninjured smooth muscle cells, since it has been suggested that altered vascular function in hypertension may be related to the resetting of the resting membrane potential in spontaneously hypertensive rats (SHR). Neointima formation was induced by balloon injury to the carotid arteries of SHR and renovascular hypertensive rats (1K-1C), as well as in their normotensive controls, i.e., Wistar Kyoto (WKY) and normal Wistar (NWR) rats. After 14 days the animals were killed and the carotid arteries were submitted to histomorphometric and immunohistochemical analyses. Resting membrane potential measurements showed that uninjured carotid arteries from SHR smooth muscle cells were significantly depolarized (-46.5 ± 1.9 mV) compared to NWR (-69 ± 1.4 mV), NWR 1K-1C (-60.8 ± 1.6 mV), WKY (-67.1 ± 3.2 mV) and WKY 1K-1C (-56.9 ± 1.2 mV). The SHR arteries responded to balloon injury with an enhanced neointima formation (neo/media = 3.97 ± 0.86) when compared to arteries of all the other groups (NWR 0.93 ± 0.65, NWR 1K-1C 1.24 ± 0.45, WKY 1.22 ± 0.32, WKY 1K-1C 1.15 ± 0.74). Our results indicate that the increased fibroproliferative response observed in SHR is not related to the hypertensive state but could be associated with the resetting of the carotid smooth muscle cell resting membrane potential to a more depolarized state.

HHV-8 infection in patients with AIDS-related Kaposi's sarcoma in Brazil

The aims of the present study were to determine the prevalence of human herpesvirus type 8 (HHV-8) in HIV-positive Brazilian patients with (HIV+/KS+) and without Kaposi's sarcoma (HIV+/KS-) using PCR and immunofluorescence assays, to assess its association with KS disease, to evaluate the performance of these tests in detecting HHV-8 infection, and to investigate the association between anti-HHV-8 antibody titers, CD4 counts and staging of KS disease. Blood samples from 66 patients, 39 HIV+/KS+ and 27 HIV+/KS-, were analyzed for HHV-8 viremia in peripheral blood mononuclear cells by PCR and HHV-8 antigenemia for latent and lytic infection by immunofluorescence assay. Positive samples for latent nuclear HHV-8 antigen (LNA) antibodies were titrated out from 1/100 to 1/409,600 dilution. Clinical information was collected from medical records and risk behavior was assessed through an interview. HHV-8 DNA sequences were detected by PCR in 74.3% of KS+ patients and in 3.7% of KS- patients. Serological assays were similar in detecting anti-LNA antibodies and anti-lytic antigens in sera from KS+ patients (79.5%) and KS- patients (18.5%). HHV-8 was associated with KS whatever the method used, i.e., PCR (odds ratio (OR) = 7.4, 95% confidence interval (CI) = 2.16-25.61) or anti-LNA and anti-lytic antibodies (OR = 17.0, 95%CI = 4.91-59.14). Among KS+ patients, HHV-8 titration levels correlated positively with CD4 counts (rho 0.48, P = 0.02), but not with KS staging. HHV-8 is involved in the development of KS in different geographic areas worldwide, as it is in Brazil, where HHV-8 is more frequent among HIV+ patients. KS severity was associated with immunodeficiency, but no correlation was found between HHV-8 antibody titers and KS staging.

Gender-related differences in the effects of nitric oxide donors on neuroleptic-induced catalepsy in mice

It has been suggested that nigrostriatal dopaminergic transmission is modulated by nitric oxide (NO). Since there is evidence that gonadal hormones can affect extrapyramidal motor behavior in mammals, we investigated the effects of isosorbide dinitrate (ISD), linsidomine (SIN-1) and S-nitroso-N-acetylpenicillamine (SNAP), three pharmacologically different NO donors, on neuroleptic-induced catalepsy in 60- to 80-day-old male and female albino mice. Catalepsy was induced with haloperidol (1 mg/kg, ip) and measured at 30-min intervals by means of a bar test. Drugs (or appropriate vehicle) were injected ip 30 min before haloperidol, with each animal being used only once. ISD (5, 20 and 50 mg/kg) caused a dose-dependent inhibition of catalepsy in male mice (maximal effect 120 min after haloperidol: 64% inhibition). In the females only at the highest dose of ISD was an attenuation of catalepsy observed, which was mild and short lasting. SIN-1 (10 and 50 mg/kg) did not significantly affect catalepsy in female mice, while a significant attenuation was observed in males at the dose of 50 mg/kg (maximal inhibition: 60%). SNAP (20 mg/kg) significantly attenuated catalepsy in males 120 min after haloperidol (44% inhibition), but had no significant effect on females. These results basically agree with literature data showing that NO facilitates central dopaminergic transmission, although the mechanisms are not fully understood. They also reveal the existence of gender-related differences in this nitrergic modulation in mice, with females being less affected than males.

Postprandial symptoms in dysmotility-like functional dyspepsia are not related to disturbances of gastric myoelectrical activity

Gastric dysrhythmias, such as tachy- or bradygastria, have been reported in patients with functional dyspepsia (FD), but their role in symptom production is uncertain. It is also not known whether gastric dysrhythmias in these patients can be elicited by physiological gastric distension with a meal. We investigated the relationships between symptoms after ingestion of different volumes of water following a test meal and gastric dysrhythmias in FD patients. Fourteen patients with dysmotility-like FD and 13 healthy volunteers underwent paired electrogastrography (EGG) studies. Fasted subjects ingested 150 ml of yoghurt with either 150 ml (low volume) or 300 ml (high volume) water in random order. Fasting and fed EGGs with monitoring of symptoms were performed in both studies. Ten FD patients (71.4%) reported upper abdominal discomfort and bloating after the low volume meal, but only one (7.1%) presented an abnormal EGG (dominant frequency in the 2-4-cpm range: 58%). Following the high volume meal, 7 patients (50%) had symptoms, but none had EGG abnormalities. No significant differences were found between FD patients and controls for any of the EGG variables, in any test. In FD patients with postprandial symptoms, the percentage of the EGG dominant frequency in the normal range (median, 84.6%; range, 76.0-100.0%) was similar (P > 0.20) to that in those without symptoms (88.5%; 75.0-100.0%). We conclude that disturbances of gastric myoelectrical activity are unlikely to play a role in the origin of postprandial upper abdominal discomfort and bloating in dysmotility-like FD.

Estrogen and progesterone receptors in human papilloma virus-related cervical neoplasia

Estrogen (ER) and progesterone (PR) receptors in the normal uterine cervix, cervical intraepithelial neoplasia and invasive carcinoma were studied in consecutive samples from Hospital do Câncer, São Paulo, between 1996 and 1997. Tissue was collected by removing a fragment of the tumoral area using a 5-mm diameter biopsy punch, followed by removal of a macroscopically normal area as close as possible from the tumor. Histopathological confirmation was obtained for all specimens analyzed. A total of 24 normal tissues, 17 cases of cervical intraepithelial neoplasia and 7 of invasive carcinomas were studied. The ER/PR ratio was determined by immunohistochemistry using monoclonal antibodies specific for each receptor. Adjacent tissue slides were submitted to generic PCR for human papillomavirus (HPV) DNA detection followed by typing by dot blot hybridization. About half (45.8%) of the tumors were HPV DNA positive while 29.1% of the patients were also HPV positive in their respective normal tissue. ER was negative in the tumoral epithelium of 11 HPV-positive patients (P = 0.04). There was a trend in the ER distribution in normal tissue that was opposite to that from lesions, but it was not statistically significant (P = 0.069). No difference in ER distribution in stromal tissues was observed between HPV-positive and HPV-negative tissues. PR staining was negative in the epithelium of all cases studied. The results obtained from this small number of cases cannot be considered to be conclusive but do suggest that factors related to viral infection affect the expression of these ER/PR cervix receptors.

Curcumin induces human HT-29 colon adenocarcinoma cell apoptosis by activating p53 and regulating apoptosis-related protein expression

Curcumin, a major yellow pigment and active component of turmeric, has multiple anti-cancer properties. However, its molecular targets and mechanisms of action on human colon adenocarcinoma cells are unknown. In the present study, we examined the effects of curcumin on the proliferation of human colon adenocarcinoma HT-29 cells by the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide method and confirmed the curcumin-induced apoptosis by morphology and DNA ladder formation. At the same time, p53, phospho-p53 (Ser15), and other apoptosis-related proteins such as Bax, Bcl-2, Bcl-xL, pro-caspase-3, and pro-caspase-9 were determined by Western blot analysis. The colon adenocarcinoma cells were treated with curcumin (0-75 µM) for 0-24 h. We observed that p53 was highly expressed in HT-29 cells and curcumin could up-regulate the serine phosphorylation of p53 in a time- and concentration-dependent manner. An increase in expression of the pro-apoptotic factor Bax and a decrease in expression of the anti-apoptotic factor Bcl-2 were also observed in a time-dependent manner after exposure of 50 µM curcumin, while the expression of the anti-apoptotic factor Bcl-xL was unchanged. Curcumin could also down-regulate the expression of pro-caspase-3 and pro-caspase-9 in a time-dependent manner. These data suggest a possible underlying molecular mechanism whereby curcumin could induce the apoptosis signaling pathway in human HT-29 colon adenocarcinoma cells by p53 activation and by the regulation of apoptosis-related proteins. This property of curcumin suggests that it could have a possible therapeutic potential in colon adenocarcinoma patients.

p16INK4 expression in precursor lesions of squamous cell cervical cancer related to the presence of HPV-DNA

The purpose of the present study was to identify the expression of p16INK4 in cervical cancer precursor lesions by immunohistochemistry and to correlate it with lesion grade and presence of human papillomavirus (HPV) infection. Cervical specimens from 144 women seen consecutively at the gynecology outpatient clinic of our institution from December 2003 to May 2005 were analyzed by cytopathology, histopathology, polymerase chain reaction for HPV-DNA, and p16INK4 immunostaining. Histologically normal biopsies, HPV-DNA negative by polymerase chain reaction, were used as control. HPV-DNA prevalence, including the control group, was 68.1% and the prevalence of p16INK4 expression was 55.0%. The percentage of cells stained by p16INK4 ranged from 10 to 100%, both in the group consisting of cervical intraepithelial neoplasia (CIN)1/HPV specimens and in the group of CIN2/CIN3 specimens with P value of 0.0001. p16INK4 expression was 48.3% in the CIN1/HPV group, as opposed to 94.3% in the CIN2/CIN3 group (P = 0.001), showing a statistically significant difference between the two groups. The quantitative method used here is simple and less subjective than the different semiquantitative methods described in the literature. In view of the different definitions of a p16INK4-positive case, it is almost impossible to compare the findings reported by different investigators. This study confirms the association between p16INK4 and CIN2 and CIN3 lesions. Moreover, it shows that some low grade lesions expressed high levels of this protein. This may indicate that such low grade lesions may be predisposed to progress to high grade lesions. This means that p16INK4 may be a strong marker for "neoplastic lesions" induced by HPV and not just an infection marker.

Tissue expression and subcellular localization of Mipu1, a novel myocardial ischemia-related gene

Myocardial ischemic preconditioning up-regulated protein 1 (Mipu1), a novel zinc finger protein, was originally cloned using bioinformatic analysis and 5' RACE technology of rat heart after a transient myocardial ischemia/reperfusion procedure in our laboratory. In order to investigate the functions of Mipu1, the recombinant prokaryotic expression vector pQE31-Mipu1 was constructed and transformed into Escherichia coli M15(pREP4), and Mipu1-6His fusion protein was expressed and purified. The identity of the purified protein was confirmed by mass spectrometry. The molecular mass of the Mipu1 protein was 70.03779 kDa. The fusion protein was intracutaneously injected to immunize New Zealand rabbits to produce a polyclonal antibody. The antibody titer was approximately 1:16,000. The antibody was tested by Western blotting for specificity and sensitivity. Using the antibody, it was found that Mipu1 was highly expressed in the heart and brain of rats and was localized in the nucleus of H9c2 myogenic cells. The present study lays the foundation for further study of the biological functions of Mipu1.

Effects of reversible inactivation of the dorsomedial hypothalamus on panic- and anxiety-related responses in rats

The medial hypothalamus is part of a neurobiological substrate controlling defensive behavior. It has been shown that a hypothalamic nucleus, the dorsomedial hypothalamus (DMH), is involved in the regulation of escape, a defensive behavior related to panic attacks. The role played by the DMH in the organization of conditioned fear responses, however, is less clear. In the present study, we investigated the effects of reversible inactivation of the DMH with the GABA A agonist muscimol on inhibitory avoidance acquisition and escape expression by male Wistar rats (approximately 280 g in weight) tested in the elevated T-maze (ETM). In the ETM, inhibitory avoidance, a conditioned defensive response, has been associated with generalized anxiety disorder. Results showed that intra-DMH administration of the GABA A receptor agonist muscimol inhibited escape performance, suggesting an antipanic-like effect (P < 0.05), without changing inhibitory avoidance acquisition. Although a higher dose of muscimol (1.0 nmol/0.2 µL; N = 7) also altered locomotor activity in an open field when compared to control animals (0.2 µL saline; N = 13) (P < 0.05), the lower dose (0.5 nmol/0.2 µL; N = 12) of muscimol did not cause any motor impairment. These data corroborate previous evidence suggesting that the DMH is specifically involved in the modulation of escape. Dysfunction of this regulatory mechanism may be relevant in the genesis/maintenance of panic disorder.

Effects of chronic corticosterone and imipramine administration on panic and anxiety-related responses

It is known that chronic high levels of corticosterone (CORT) enhance aversive responses such as avoidance and contextual freezing. In contrast, chronic CORT does not alter defensive behavior induced by the exposure to a predator odor. Since different defense-related responses have been associated with specific anxiety disorders found in clinical settings, the observation that chronic CORT alters some defensive behaviors but not others might be relevant to the understanding of the neurobiology of anxiety. In the present study, we investigated the effects of chronic CORT administration (through surgical implantation of a 21-day release 200 mg pellet) on avoidance acquisition and escape expression by male Wistar rats (200 g in weight at the beginning of the experiments, N = 6-10/group) tested in the elevated T-maze (ETM). These defensive behaviors have been associated with generalized anxiety and panic disorder, respectively. Since the tricyclic antidepressant imipramine is successfully used to treat both conditions, the effects of combined treatment with chronic imipramine (15 mg, ip) and CORT were also investigated. Results showed that chronic CORT facilitated avoidance performance, an anxiogenic-like effect (P < 0.05), without changing escape responses. Imipramine significantly reversed the anxiogenic effect of CORT (P < 0.05), although the drug did not exhibit anxiolytic effects by itself. Confirming previous observations, imipramine inhibited escape responses, a panicolytic-like effect. Unlike chronic CORT, imipramine also decreased locomotor activity in an open field. These data suggest that chronic CORT specifically altered ETM avoidance, a fact that should be relevant to a better understanding of the physiopathology of generalized anxiety and panic disorder.

Chronotype ontogeny related to gender

Chronotype is an established concept designed to identify distinct phase relationships between the expression of circadian rhythms and external synchronizers in humans. Although it has been widely accepted that chronotype is subjected to ontogenetic modulation, there is no consensus on the interaction between age and gender. This study aimed to determine the relationship between age- and gender-related changes in the morningness-eveningness character in a large sample of people. A total of 14,650 volunteers were asked to complete the Brazilian version of the Horne and Östberg chronotype questionnaire. The data demonstrated that, on average, women were more morning-oriented than men until the age of 30 and there were no significant differences between men and women from 30 to 45 years of age. In contrast to the situation observed until the age of 30, women older than 45 years were more evening-oriented than men. These results suggest that the ontogenetic development of the circadian timekeeping system is more plastic in men, as represented by the larger amplitude of chronotype changes throughout their aging process. The phase delay of adolescence and phase advance of the elderly seem to be phenomena that are more markedly present in men than in women. Thus, our data, for the first time, provide support that sharply opposes the view that there is a single path toward morningness as a function of age, regardless of gender.