Search PubMed⌕ Search

Biomedical subjects

P Pacheco

Publications and source records attributed to P Pacheco.

At least 37 records · Page 2Linked to original sources
PubMed1999

Bradykinin down-regulates LPS-induced eosinophil accumulation in the pleural cavity of mice through type 2-kinin receptor activation: a role for prostaglandins.

1. The role of both exogenously administered and endogenously generated bradykinin (BK) on LPS-induced eosinophil accumulation in the mice pleural cavity was investigated by means of treatment with BK selective receptor agonists/antagonists and captopril. 2. Intrathoracic (i.t.) injection of LPS (250 ng cavity(-1)) induced eosinophil influx at 24 h as previously described (Bozza et al., 1993). Pretreatment with the B1 receptor antagonist des-Arg9-[leu-8]BK (0.025 and 0.25 nmol cavity(-1)) showed no effect on this phenomenon, whereas pretreatment with the B2 receptor antagonists, NPC 17731 (0.025 and 0.25 nmol cavity(-1)) or HOE 140 (2.5 nmol cavity(-1)), increased LPS-induced eosinophil influx. Accordingly, pretreatment with captopril at 10 mg kg(-1) i.p., inhibited eosinophil infiltration induced by LPS in the pleural cavity, suggesting that endogenous BK is down-regulating LPS-induced eosinophil accumulation. 3. BK administered at 15 and 25 nmol cavity(-1), i.t. or i.p. also inhibited LPS-induced eosinophil accumulation. BK alone had no effect on the basal number of leucocytes in the pleural or peritoneal cavity in doses up to 25 nmol cavity(-1). Nevertheless, when injected at doses of 50 and 100 nmol cavity(-1) BK induced leucocyte influx characterized by neutrophil and eosinophil accumulation at 24 h. 4. Similarly to what was observed with BK, a specific B2 receptor agonist, Tyr8BK, administered at 0.25 nmol cavity(-1) i.p., significantly inhibited the eosinophil influx induced by LPS. 5. The mechanism by which B2 receptor agonists inhibit LPS-induced eosinophil accumulation was investigated by pretreating the animals with indomethacin or a selective cyclooxygenase-2 inhibitor, NS-398. Pretreatment with either indomethacin or NS-398 had no effect on eosinophil influx induced by LPS alone, but those drugs were able to restore the LPS-induced eosinophil influx in Tyr8BK (0.25 nmol cavity(-1)) injected mice. 6. In conclusion, endogenously generated bradykinin seems to modulate, through activation of B2 receptors, eosinphil accumulation induced by LPS via a mechanism dependent on prostanoid synthesis.

Angiotensin-Converting Enzyme Inhibitors↗
PubMed1999

Separate taurine and chloride efflux pathways activated during regulatory volume decrease.

Organic osmolyte and halide permeability pathways activated in epithelial HeLa cells by cell swelling were studied by radiotracer efflux techniques and single-cell volume measurements. The replacement of extracellular Cl- by anions that are more permeant through the volume-activated Cl- channel, as indicated by electrophysiological measurements, significantly decreased taurine efflux. In the presence of less-permeant anions, an increase in taurine efflux was observed. Simultaneous measurement of the 125I, used as a tracer for Cl-, and [3H]taurine efflux showed that the time courses for the two effluxes differed. In Cl--rich medium the increase in I- efflux was transient, whereas that for taurine was sustained. Osmosensitive Cl- conductance, assessed by measuring changes in cell volume, increased rapidly after hypotonic shock. The influx of taurine was able to counteract Cl- conductance-dependent cell shrinkage but only approximately 4 min after triggering cell swelling. This taurine-induced effect was blocked by DIDS. Differences in anion sensitivity, the time course of activation, and sensitivity to DIDS suggest that the main cell swelling-activated permeability pathways for taurine and Cl- are separate.

Anions↗
PubMed1998

The sensory but not muscular pelvic nerve branch is necessary for parturition in the rat.

In the rat the pelvic nerve consists of a viscerocutaneous (sensory) branch which receives information from pelvic viscera and the midline perineal region, and a somatomotor (muscular) branch which innervates the ilio- and pubococcygeous muscles. To investigate the contribution of these branches to the parturition process, the length of gestation and course of delivery were closely monitored in 43 pregnant, Wistar-strain rats randomly assigned to five groups: untreated control animals, animals in which the somatomotor branch of the pelvic nerve was bilaterally sectioned on Day 14 of gestation, animals in which the viscerocutaneous branch of the pelvic nerve was bilaterally sectioned on Day 14 of gestation, animals treated similarly to the previous group but with young delivered by C-section at term, and sham-operated controls. Sectioning the viscerocutaneous branch seriously disrupted parturition and resulted in major dystocia and a high percentage of stillbirths in all females. In contrast, sectioning the somatomotor branch had no apparent effect on parturition and no significant differences were found between females of this group and sham or control dams on any of the measures recorded. It is concluded that the viscerocutaneous branch of the pelvic nerve is vital for the normal course of parturition in the rat but that the somatomotor branch plays little role, if any.

Abdominal Muscles↗
PubMed1997

Electrophysiological evidence for the nomenclature of the pudendal nerve and sacral plexus in the male rat.

Surgical microscopy and electrophysiological techniques were used to standardize the nomenclature for the pudendal nerve and sacral plexus according to their somatic axonal composition in the male rat. We conclude that the pudendal nerve is the segment running from the L6-S1 trunk to the sacral plexus, carrying efferent fibers to the coccygeus, internal obturator, ventral and dorsal bulbospongiosus, ischiocavernosus, external anal sphincter, and external urethral sphincter muscles, and afferent fibers from the penis, prepuce, scrotum, and ventral-proximal tail. The sacral plexus is the complex formed by the bridge-like structure connecting the pudendal nerve with the lumbosacral trunk, and two nerve branches emerging from it, one innervating the proximal half of the scrotal skin, and the other innervating the muscles at the base of the penis known as the motor branch. These branches are only considered as a part of the sacral plexus because they integrate axons from both the lumbosacral trunk and pudendal nerve. The gross anatomy of the pudendal nerve and sacral plexus has a main organization that was observed in 70% of cases, whereas the remaining 30% occurred in two variants. This nomenclature is appropriate to describe the pudendal nerve and sacral plexus in studies that involve them being lesioned or electrophysiologically analysed. A main additional finding was that two large afferent branches innervate the scrotum, one the proximal half and the other the distal half. As mentioned above, the proximal branch belongs to the sacral plexus, whereas the distal branch belongs to the pudendal nerve because all its axons travel to the cord via this nerve. Since stimulation or even manipulation of the scrotal branches resulted in the secretion of semen containing spermatozoa, it is suggested that scrotal afferents are involved in some way in the ejaculatory process, a topic that deserves further research.

Animals↗
PubMed1997

The divergent 5' termini of the alpha human folate receptor (hFR) mRNAs originate from two tissue-specific promoters and alternative splicing: characterization of the alpha hFR gene structure.

The human KB cell or alpha folate receptor (alpha hFR) is a membrane glycoprotein of 42 kDa that participates in the internalization of folates and antifolates. Seven independent alpha hFR cDNA isoforms have been reported that contain unique 5' termini but share a common open reading frame (ORF). To investigate the molecular basis of these heterogeneous 5' sequences, we determined the sequence of the alpha hFR gene from two clones isolated from a human lymphocyte lambda DASH genomic library. The gene is composed of seven exons that span 6.8 kb. The ORF is encoded by exons 4 through 7 while the reported 5' termini of the cDNA isoforms (including two novel cDNAs designated KB2 and KB4) are encoded by exons 1 through 4. Using RNase protection assays, we demonstrate that transcripts corresponding to the KB1 and KB4 cDNAs originate from promoters upstream from exon 1 and exon 4, designated P1 and P4, respectively, and that these mRNA isoforms are the most abundant transcripts expressed in KB cells and selected normal tissues (including kidney, lung, and cerebellum). We observed a heterogeneous start site within exon 1 from the P1 promoter while transcripts from the P4 promoter originate from a single site. In addition, we detected tissue specificity for the P1 and P4 promoter utilization. Transcripts originating from the P1 promoter are the most abundant transcripts expressed by human cerebellum and kidney. In contrast, transcripts from the P4 promoter are the most abundant transcripts expressed by human KB cells and lung. Total RNA from KB cells also protects a 66 bp fragment of an exon 3 riboprobe that is consistent with an alternatively spliced transcript. To examine the functional activity of the predicted P1 and P4 promoters, alpha hFR promoter-CAT chimeric plasmids were constructed using sequences flanking exon 1 and exon 4. We observed a 7.5- and 10-fold increase in CAT activity in HeLa cells transiently transfected with the P1 and P4 promoter constructs, respectively. These data demonstrate that a single gene encodes the divergent 5' termini of the alpha hFR cDNAs and that the alpha hFR transcripts are transcribed from two promoters that are activated in a tissue-specific manner.

Alternative Splicing↗
PubMed1997

Striated muscles and scent glands associated with the vaginal tract of the rabbit.

BACKGROUND: Despite the importance of the rabbit (Oryctolagus cuniculus) in reproductive research, no comprehensive account of the anatomical organization of the female reproductive tract is presently available. METHODS: Based on gross dissection and on the electrical stimulation of striated muscles in 10 domestic female rabbits, it was the aim of this study to help fill this gap by describing the origin, insertion, and general action of muscles associated with the vaginal tract and with associated scent glands. RESULTS: Compared with the better-studied rat or cat, the long vaginal tract of the rabbit is characterized by a well-developed and complex musculature. The bulbospongiosus, ischiocavernosus, constrictor vestibuli, and constrictor vulvae muscles, poorly developed or absent in the female rat or cat, were particularly conspicuous. Scent glands, including the newly identified preputial glands, were also a distinctive feature. CONCLUSION: These structures may represent specializations underlying the rapid copulation, rapid parturition, and well-developed chemical communication characteristic of the reproductive efficiency of the rabbit.

Animals↗
PubMed1997

Missense mutation R1066C in the second transmembrane domain of CFTR causes a severe cystic fibrosis phenotype: study of 19 heterozygous and 2 homozygous patients.

We report the clinical features of 21 unrelated cystic fibrosis (CF) patients from Portugal and Spain, who carry the mutation R1066C in the CFTR gene. The current age of the patients was higher in the R1066C/any mutation group (P < 0.01), as compared to the deltaF508/deltaF508 group. Poor values for lung radiological involvement (Chrispin-Norman) and general status (Shwachman-Kulcycki) were observed in the R1066C/any mutation group (P < 0.005 and P < 0.0004). A slightly, but not significantly worse lung function was found in the R1066C/any mutation group when compared with the deltaF508/deltaF508 patients. No significant differences were detected regarding the age at diagnosis, sweat Cl-values, or percentiles of height and weight between the two groups. Neither were significant differences observed regarding sex, meconium ileus (4.7% vs. 11.1%), dehydration (10.5% vs. 14.7%), or pancreatic insufficiency (PI) (100% vs. 97.8%). The proportion of patients with lung colonization by bacterial pathogens was slightly, but not significantly higher in the R1066C/any mutation group (70.0%), as compared with the deltaF508/deltaF508 group (57.5%). Other clinical complications were significantly more frequent in the R1066C/any mutation patients(P < 0.02) than in the deltaF508/deltaF508 group. The two homozygous R1066C/R1066C patients died at the ages of 3 months and 7 years. The data presented in this study clearly demonstrate that the R1066C mutation is responsible for a severe phenotype similar to that observed in homozygous deltaF508 patients. The poor clinical scores and complications of patients with the R1066C mutation are probably related to their slightly longer survival.

Adolescent↗
PubMed1997

The development of auditory callosal connections in normal and hypothyroid rats.

Previous studies have shown that hypothyroidism modifies the development of callosal connections. In particular, adult hypothyroid rats have fewer callosally projecting neurons in layers II-III of the auditory cortex and more in layer V. This might be due to disturbance in the stabilization/elimination of juvenile callosal axons, or to abnormal neuronal migration during cortical histogenesis. To distinguish between these possibilities we have studied the distribution of callosally projecting auditory neurons at different postnatal ages using retrogradely transported tracers, and the cortical neurogenetic gradients using DNA labelling with 5-bromo-2'-deoxiuridine. In hypothyroid rats, injected at postnatal day 5 (P5) and killed at P18-20, most of the neurons retrogradely labelled from the contralateral hemisphere are distributed between layers IV and VI, as in older rats. In hypothyroid rats, many neurons are at locations inappropriate for their birthdate, including the subcortical white matter, resulting in more diffuse radial neurogenetic gradients. These results indicate that early induced hypothyroidism alters neuronal migration and prevents the establishment of callosal connections from cortical layers II-III.

Animals↗
PubMed1997

The role of pubococcygeus muscle in urinary continence in the male rat.

The role of the male rat pubococcygeus muscle (Pcm) in the micturition reflex was analyzed. Anatomical features of Pcm, electrical stimulation of its nerve, electrical recording and stimulation of the muscle and cystometrograms were carried out. Results showed that Pcm has fibers attached to the ventrolateral part of the external urethral sphincter, and that its activity contributes to hold the tail in the midline. Pcm shows activity during fluid expulsion in cystometrograms and spontaneous micturitions. This activity produced reflex inhibition of detrusor contraction and was not the cause of intravesical high frequency oscillations. Thus, it is proposed that Pcm activity produces the discharge of its afferents which in turn activates a spinal reflex to promote continence.

Animals↗
PubMed1996

Complex cystic fibrosis allele R334W-R1158X results in reduced levels of correctly processed mRNA in a pancreatic sufficient patient.

CFTR alleles containing two mutations have been very rarely found in cystic fibrosis (CF) patients. They provide an opportunity to study the effect of two in cis-interacting gene defects on gene expression. Here, we describe a three-generation CF family with a complex CFTR allele that has not been previously described, containing the missense mutation R334W in exon 7 and the nonsense mutation R1158X in exon 19. Lymphocyte RNA analysis showed that (1) the mRNA corresponding to the complex allele is present although at markedly reduced levels; and (2) the nonsense mutation does not lead to detectable skipping of exon 19. The clinical picture of the patients with the genotype R334W-R1158X/delta F508 is characterized by pancreatic sufficiency and an atypical course of the disease.

Alleles↗
PubMed1996

Changes in pain threshold during the reproductive cycle of the female rat.

Responsiveness to pain was determined in female rats across the whole reproductive cycle using the tail-flick test. When tested immediately after mating, pain thresholds were unaltered, whereas 10 min later animals typically demonstrated hyperalgesia (Experiment 1). They also demonstrated hyperalgesia during most of pregnancy, and had significantly lower pain thresholds than the unmated controls except for the 24 h before parturition, when a sudden increase in tail-flick latencies was recorded (Experiment 2). Pain thresholds were also significantly lower throughout the nursing period but increased significantly when dams were separated from their litters for 6 h, and returned to premating baseline values within 24 h of weaning (Experiment 3). These findings confirm and extend earlier reports that female reproductive state may significantly modify responsiveness to noxious stimuli, and it is suggested that differences between the results of this and previous studies may be at least partly explained by the relatively stress-free test procedure adopted here.

Animals↗
PubMed1996

Delayed expression of the insulin-like growth factor I (IGF-I) gene in the XY sex-reversed female mouse ovary.

When the Y chromosome of Mus musculus domesticus (YDOM) mouse strain from Tirano (Italy) or Mus musculus poschiavinus (YPOS) from Poschiavo (Switzerland), is placed onto the C57BL/6J (B6) inbred background, the YDOM chromosome fails to induce normal testicular differentiation and instead allows development of ovaries and ovotestes in embryonic life. During postnatal development some hermaphroditic males become fertile whereas the XY females lack normal estrus cyclicity, produce low levels of gonadal steroids and cannot carry pregnancy to term. Here we studied the transcription of the IGF-I gene known to be involved in steroidogenesis. RNA was isolated from the XX and the XY ovaries at 1 to 40 d.p.p. and subjected to RT-PCR analysis. Immunocytochemical staining for IGF-I was performed to identify the cell type of IGF-I peptide localization, and protein expression was examined by Western blot analysis. The present results indicate that the IGF-I transcript was expressed at 1 d.p.p. in the XX ovary throughout the studied stages whereas in the XY ovary mRNA IGF-I was not detected until 15 d.p.p. IGF-I protein was identified in theca cells in the whole XX control ovary, while in the XY ovary, strong staining for IGF-I was found in the theca cells of the cortex. Faint staining was also seen around the medullary sterile cords. Western blot analysis showed normal onset in the XX and the XY ovary, but a different staining pattern for IGF-I in the XY ovary at 11 and 26 d.p.p. compared to the XX control ovary. We propose that delayed expression of IGF-I in the XY mouse ovary may be responsible for low steroid production and fertility problems in the XY sex-reversed adult female mouse.

Animals↗
PubMed1995

Brain 2-deoxyglucose levels related to maternal behavior-inducing stimuli in the rat.

Levels of [14C]2-deoxyglucose (2-DG), measured autoradiographically, in the medial preoptic area (MPOA), were higher during natural parturition with concurrent maternal behavior than in non-pregnant non-maternal controls, whereas levels in the vomeronasal system were lower in virgin rats made maternal by cohabitation with young than in control and parturient rats. Previous studies have shown that lesions of MPOA disrupt maternal behavior, whereas lesions of vomeronasal structures stimulate it, and that an increase in 2-DG levels is indicative of an increase in firing activity in neuron terminals. Consequently, the present findings suggest that maternal behavior can be induced by: (a) an increase in parturition-generated sensory stimulatory input to the MPOA in response to mechanostimulation of the birth canal, and (b) a separate chemosensory vomeronasal pathway whose activity is reduced cohabitation with young, thereby disinhibiting maternal behavior.

Animals↗
PubMed1995

Beta-thalassaemia unlinked to the beta-globin gene interacts with sickle-cell trait in a Portuguese family.

An autosomally transmitted hypochromic microcytic mild anaemia with elevated haemoglobin (Hb) A2 and globin chain imbalance has been observed in a three-generation family of Portuguese origin. Extensive DNA analysis of the beta-globin gene cluster, including the complete sequencing of the beta-globin gene and flanking regions, failed to reveal any genetic alteration. The co-segregation of sickle-cell trait in this family enabled us to postulate a defective erythroid trans-acting factor was playing a role in the down-regulation of both beta A- and beta S-globin genes. Among the transcription factors that could possibly have caused the reported phenotype, NF-E2 is unlikely to be implicated, whereas Nrf1 and Nrf2 cannot be ruled out. Thus, this family carries a novel beta-thalassaemia autosomal determinant unlinked to the beta-globin gene. This observation reinforces the notion of the haemoglobinopathies as single gene disorders under polygenic regulation.

Adult↗
PubMed1995

Central effects of catecholamines upon mammary contractility in rats are neurally mediated.

We injected, i.e., intracerebroventricularly (ICV) or systematically, small amounts of adrenaline (ADR), noradrenaline (NA), isoproterenol (ISOP) and dopamine (DA) in urethane-anesthetized lactating rats, and determined the effects on isometrically recorded intramammary pressure (IMP) responses to exogenous oxytocin (OXY). While centrally administered ADR, NA and DA provoked increased IMP responses to OXY, the beta-adrenergic agonist ISOP induced the opposite effect. These effects were reversible, dose related and also occurred in hypophysectomized rats. However, when injected systematically, all adrenergic agonists but DA depressed IMP responses to OXY. Further experiments showed that central effects of catecholamines were exerted by regulating ductal tone, through the direct innervation of the mammary glands. Thus, whereas complete blockage of these effects occurred after selective denervation of the mammary glands, increased ductal tone resulted from ICV administration of ISOP. Finally, evidence was also obtained that antagonistic alpha- and beta-adrenergic mechanisms may interact with each other to regulate milk ejection, and with afferent signals from the mammary glands. Thus, beta-adrenergic inhibition upon IMP was counteracted by either NA administration or by activation of ductal mechanoreceptors. Together, these results suggest that regulation of milk ejection may involve neurally mediated influences on mammary contractility. Such actions would interact closely with afferents from the mammary gland influencing ductal tone.

Adrenergic Agents↗
PubMed1995

Neuroendocrine regulation of adrenal gland and hypothalamus 5'deiodinase activity. II. Effects of splanchnicotomy and hypophysectomy.

This study analyzes the role of the autonomic nervous system, the pituitary gland, ACTH, dexamethasone (DEX), and thyroid hormones in the regulation of 5'deiodinase (5'D) in the hypothalamus (HP) and adrenal gland (AG) of the rat. 5'D activity was analyzed in rats under basal conditions (22 C) and during cold exposure (4 C, during 15, 30, 60, and 120 min). Several experimental groups were formed: intact animals (INT), unilateral (left) splanchnicotomized, sham splanchnicotomized, hypophysectomized (HPX), and sham hypophysectomized. Results in the hypothalamus were: 1) independent of the experimental group, the HP 5'D activity increased during the first 15-30 min of cold exposure; however, this increase was greater in operated animals than in INT rats; and 2) basal 5'D activity was increased in HPX rats and was also regulated by thyroid hormones. Results in the adrenal gland were: 1) INT rats exhibited a biphasic pattern of 5'D activation during cold stress (30 and 60 min of exposure); 2) the splanchnic nerve exerted a tonic-stimulatory effect on basal AG 5'D activity; 3) the denervated gland preserved its ability to respond to cold; 4) in INT animals DEX but not ACTH had a stimulatory effect on basal activity; 5) the high 5'D activity post-HPX was reverted to basal values by T4 and DEX administration; 6) SHAM-HPX also was followed by a large increase in basal 5'D activity, and 7) this hyperresponse was abolished by acute ACTH and DEX administration. In conclusion, our results demonstrate that the mechanisms that participate in the regulation and activation of 5'D in the adrenal gland and the hypothalamus are of a neuroendocrine nature. Also, in both organs, but mainly in the HP, 5'D activity is T4-dependent. In addition to the tonic-stimulatory influence conveyed by the splanchnic nerve, AG 5'D activity is influenced by thyroid hormones, glucocorticoids, and probably extrapituitary factors whose nature is unknown yet.

Adrenal Glands↗