Paraventricular nucleus mediates pressor response to noradrenaline injection into the dorsal periaqueductal gray area

The dorsal periaqueductal gray area (dPAG) is involved in cardiovascular modulation. In a previous study, we reported that noradrenaline (NA) microinjection into the dPAG of rats caused pressor response that was mediated by vasopressin release. Vasopressin is synthesized by magnocellular neurons in the hypothalamic paraventricular (PVN) and supraoptic (SON) nuclei. In the present study, we verified which nuclei mediated the cardiovascular response to NA as well as the existence of direct neural projection from the dPAG to hypothalamic nuclei. Then, we studied the effect of treating either PVN or SON with the nonselective synaptic blocker cobalt chloride (1 mM) on the cardiovascular response to NA (15 nmol) microinjection into dPAG. Attempting to identify neural projections from dPAG to hypothalamic nuclei, we microinjected the neuronal tracer biotinylated-dextran-amine (BDA) into the dPAG and searched varicosity-containing nerve terminals in the PVN and SON. Unilateral cobalt-induced inhibition of synapses in the SON did not affect the cardiovascular response to NA. However, unilateral inhibition of PVN significantly reduced the pressor response to NA. Moreover, cobalt-induced inhibition of synapses in both PVN blocked the pressor response caused by NA microinjected into the dPAG. Microinjection of BDA into the dPAG evidenced presence of varicosity-containing neuronal fibers in PVN but not in SON. The results from cobalt treatment indicated that synapses in PVN mediate the vasopressin-induced pressor response caused by NA microinjection into the dPAG. In addition, the neuroanatomical results from BDA microinjection into the dPAG pointed out the existence of direct neural projections from the dPAG site to the PVN. (C) 2009 Elsevier B.V. All rights reserved.

Modulation of arterial pressure by P2 purinoceptors in the paraventricular nucleus of the hypothalamus of awake rats

In the present study we evaluated the role of purinergic mechanisms in the PVN on the tonic modulation of the autonomic function to the cardiovascular system as well on the cardiovascular responses to peripheral chemoreflex activation in awake rats Guide-cannulae were bilaterally Implanted in the direction of the PVN of male Wistar rats Femoral artery and vein were catheterized one day before the experiments Chemoreflex was activated with KCN (30 mu g/0 05 ml iv) before and after microinjections of P2 receptors antagonist into the PVN Microinjection of PPADS a non selective P2X antagonist Into the PVN (n = 6) produced a significant increase in the baseline MAP (99 +/- 2 vs 112 +/- 3 mmHg) and HR (332 +/- 8 vs 375 +/- 8 bpm) but had no effect on the pressor and bradycardic responses to chemoreflex activation Intravenous injection of vasopres in receptors antagonist after microinjection of PPADS into the PVN produced no effect on the increased baseline MAP Simultaneous microinjection of PPADS and KYN into the PVN (n=6) had no effect in the baseline MAP HR or in the pressor and bradycardic responses to chemoreflex activation We conclude that P2 purinoceptors in the PVN are involved in the modulation of baseline autonomic function to the cardiovascular system but not in the cardiovascular responses to chemoreflex activation in awake rats (C) 2010 Elsevier B V All rights reserved

Dopamine microinjected into brainstem of awake rats affects baseline arterial pressure but not chemoreflex responses

Dopamine (DA) is a neuromodulator in the brainstem involved with the generation and modulation of the autonomic and respiratory activities. Here we evaluated the effect of microinjection of DA intracistema magna (icm) or into the caudal nucleus tractus solitarii (cNTS) on the baseline cardiovascular and respiratory parameters and on the cardiovascular and respiratory responses to chemoreflex activation in awake rats. Guide cannulas were implanted in cisterna magna or cNTS and femoral artery and vein were catheterized. Respiratory frequency (f(R)) was measured by whole-body plethysmography. Chemoreflex was activated with KCN (iv) before and after microinjection of DA icm or into the cNTS bilaterally while mean arterial pressure (MAP), heart rate (HR) and f(R) were recorded. Microinjection of DA icm (n = 13), but not into the cNTS (n = 8) produced a significant decrease in baseline MAP (-15 +/- 1 vs 1 +/- 1 mm Hg) and HR (-55 +/- 11 vs -11 +/- 17 bpm) in relation to control (saline with ascorbic acid, n = 9) but no significant changes in baseline f(R). Microinjection of DA icm or into the cNTS produced no significant changes in the pressor, bradycardic and tachypneic responses to chemoreflex activation. These data show that a) DA icm affects baseline cardiovascular regulation, but not baseline f(R) and autonomic and respiratory components of chemoreflex and b) DA into the cNTS does not affect either the autonomic activity to the cardiovascular system or the autonomic and respiratory responses of chemoreflex activation. (C) 2010 Elsevier B.V. All rights reserved.

Fos expression in the NTS in response to peripheral chemoreflex activation in awake rats

Chemoreflex afferent fibers terminate in the nucleus tractus solitarii (NTS), but the specific location of the NTS neurons excited by peripheral chemoreflex activation remains to be characterized. Here, the topographic distribution of chemoreflex sensitive cells at the commissural NTS was evaluated. To reach this goal, Fos-immunoreactive neurons (Fos-ir) were accounted in rostro-caudal levels of the intermediate and caudal commissural NTS, after intermittent chemoreflex activation with intravenous injection of potassium cyanide [KCN (80 mu g/kg) or saline (0.9%, vehicle), one injection every 3 min during 30 min]. In response to intermittent intravenous injections of KCN, a significant increase in the number of Fos-ir neurons was observed specifically in the lateral intermediate commissural NTS [(LI)NTS (82 +/- 9 vs. 174 +/- 16, cell number mean per section)] and lateral caudal commissural NTS [(LI)NTS (71 +/- 9 vs. 199 +/- 18, cell number mean per section)]. To evaluate the influence of baroreceptor-mediated inputs following the increase in blood pressure during intermittent chemoreflex activation, we performed an intermittent activation of the arterial baroreflex by intravenous injection of phenylephrine [1.5 mu g/kg iv (one injection every 3 min during 30 min)]. This procedure induced no change in Fos-ir in (LI)NTS (64 +/- 6 vs. 62 +/- 12, cell number mean per section) or (LC)NTS (56 +/- 15 vs. 77 +/- 12, cell number mean per section). These data support the involvement of the commissural NTS in the processing of peripheral chemoreflex, and provide a detailed characterization of the topographical distribution of activated neurons within this brain region. (C) 2009 Elsevier B.V. All rights reserved.

GABA and nitric oxide in the PVN are involved in arterial pressure control but not in the chemoreflex responses in rats

GABAergic, nitrergic and glutamatergic mechanisms in the PVN on the baseline mean arterial pressure (MAP), heart rate (HR) and on the cardiovascular responses to chemoreflex activation in awake rat were evaluated. Chemoreflex was activated with KCN before and after microinjections into the PVN. Bicuculline into the PVN increased baseline MAP (94+/-3 vs 113+/-5 mmHg) and HR (350+/-9 vs 439+/-18 bpm) but had no effect on the pressor (49+/-5 vs 47+/-6 mmHg) or bradicardic (-213+/-23 vs -256+/-42 bpm) responses (n=7). Kynurenic acid into the PVN (n=6) produced no significant changes in the MAP (98+/-3 vs 100+/-3 mmHg), HR (330+/-5 vs 339+/-12 mmHg) or in the pressor (50+/-4 vs 42+/-4 mmHg) and bradicardic (-252+/-4 vs -285+/-16 bpm) responses to chemoreflex. L-NAME into the PVN (n=8) produced increase in the MAP (94+/-3 vs 113+/-5 mmHg) and HR (350+/-9 vs 439+/-18 bpm) but had no effect on the pressor (52+/-5 vs 47+/-6 mmHg) or bradicardic (-253+/-19 vs -320+/-25 bpm) responses to chemoreflex. We conclude that GABA(A) and nitric oxide in the PVN are involved in the maintenance of the baseline MAP but not in the modulation of the responses to chemoreflex. The results also show that Glutamate receptors in the PVN are not involved in maintenance of the baseline MAP, HR or in the cardiovascular responses to chemoreflex in awake rats. (C) 2008 Elsevier B.V. All rights reserved.

Development and characterization of microsatellite loci in the marsh deer (Blastocerus dichotomus Cervidae)

Marsh deer (Blastocerus dichotomus) is one of the most exposed large mammals in South America. To aid in the conservation management of the species, nine polymorphic microsatellite loci were isolated and tested on up to 50 animals, showing 3-12 alleles and expected heterozygosity values varying from 0.69 to 0.89. These markers should be of considerable utility in future population and ecological genetics studies of this species. The marsh deer (Blastocerus dichotomus) is the biggest South American species of deer. Originally distributed across a large part of South America, stretching from the south bank of the Amazon river to northern Argentina, significant wild populations are now restricted to the Pantanal, swamplands that cover about 40% of southwest Brazil. The marsh deer is listed as Vulnerable on the Red List of the IUCN. Three populations of the species from three areas in the Parana River basin (between the states of Sao Paulo and Mato Grosso do Sul) were recently studied by observing protein polymorphism at 17 loci (Oliveira et al. 2005). Now we are presenting data about isolation of microsatellite markers to improve the results regarding population structure.

MHM assay: molecular sexing based on the sex-specific methylation pattern of the MHM region in chickens

Bird sex determination using molecular methods has proved to be a valuable tool in different studies. Although it is possible to sex most birds by coupling the CHD assay with others available methods, no sex-determining gene like SRY in mammalians has been identified in birds. The male hypermethylated (MHM) region on the Z chromosome has been found to be hypermethylated in males and hypomethylated in females in birds of the order Galliformes. We analyzed the DNA from feathers of 50 adult chickens to verify the methylation pattern of the MHM region by PCR and the restriction enzyme HpaII (a method named MHM assay). The results, visualized in agarose gel, were compared with PCR amplification of the CHD-Z and CHD-W genes (polyacrylamide gel) and with the birds` phenotype. All males (25) showed hypermethylation of the MHM region, and all females (25) showed hypomethylation. The sexing by MHM assay was in according with phenotype and CHD sexing. To our knowledge, this is the first study that uses the MHM region for sexing birds. Although the real role of the MHM region in the sex determination is still unclear, this could be a universal marker for sexing birds and may be involved in sex determination by its influence on transcriptional processes. The MHM assay could be a good alternative for CHD assay in developmental studies.

The new craft skills of engineering: The impact of innovation technology on engineering practices

This chapter explores the impact of innovation technologies such as simulation, modelling, and rapid prototyping on engineering practice. Innovation technologies help redefine the role of engineers in the innovation process, creating a new division of innovative labour both with and across organizations. This chapter also explores the boundaries of experimentation and inertia within particular domains of problem-solving to create new opportunities and value.

Interaction of purinergic and nitrergic mechanisms in the caudal nucleus tractus solitarii of rats

The interaction of purinergic and nitrergic mechanisms was evaluated in the caudal nucleus tractus solitarii (cNTS) using awake animals and brainstem slices. In awake animals, ATP (1.25 nmol/50 nL) was microinjected into the cNTS before and after the microinjection of a selective neuronal nitric oxide synthase (nNOS) inhibitor N-propyl-L-arginine (NPLA, 3 pmoles/50 nL, n=8) or vehicle (saline, n=4), and cardiovascular and ventilatory parameters were recorded. In brainstem slices from a distinct group of rats, the effects of ATP on the NO concentration in the cNTS using the fluorescent dye DAF-2 DA were evaluated. For this purpose brainstem slices (150 pm) containing the cNTS were pre-incubated with ATP (500 mu M; n=8) before and during DAF-2 DA loading. Microinjection of ATP into the cNTS increases the arterial pressure (AP), respiratory frequency (f(R)) and minute ventilation (V(E)), which were significantly reduced by pretreatment with N-PLA, a selective nNOS inhibitor (AP: 39 +/- 3 vs 16 +/- 14 mm Hg; f(R): 75 +/- 14 vs 4 +/- 3 cpm; V(E): 909 159 vs 77 39 mL kg(-1) m(-1)). The effects of ATP in the cNTS were not affected by microinjection of saline. ATP significantly increased the NO fluorescence in the cNTS (62 +/- 7 vs 101 +/- 10 AU). The data show that in the cNTS: a) the NO production is increased by ATP; b) NO formation by nNOS is involved in the cardiovascular and ventilatory responses to microinjection of ATP. Taken together, these data suggest an interaction of purinergic and nitrergic mechanisms in the cNTS. (C) 2009 Elsevier B.V. All rights reserved.

Nitric oxide synthesis blockade reduced the baroreflex sensitivity in trained rats

Objective: The present study has investigated the effect of blockade of nitric oxide synthesis on cardiovascular autonomic adaptations induced by aerobic physical training using different approaches: 1) double blockade with methylatropine and propranolol; 2) systolic arterial pressure (SAP) and heart rate variability (HRV) by means of spectral analysis; and 3) baroreflex sensitivity. Methods: Male Wistar rats were divided into four groups: sedentary rats (SR); sedentary rats treated with N(omega)-nitro-L-arginine methyl ester (L-NAME) for one week (SRL); rats trained for eight weeks (TR); and rats trained for eight weeks and treated with L-NAME in the last week (TRL). Results: Hypertension and tachycardia were observed in SRL group. Previous physical training attenuated the hypertension in L-NAME-treated rats. Bradycardia was seen in TR and TRL groups, although such a condition was more prominent in the latter. All trained rats had lower intrinsic heart rates. Pharmacological evaluation of cardiac autonomic tonus showed sympathetic predominance in SRL group, differently than other groups. Spectral analysis of HRV showed smaller low frequency oscillations (LF: 0.2-0.75 Hz) in SRL group compared to other groups. Rats treated with L-NAME presented greater LF oscillations in the SAP compared to non-treated rats, but oscillations were found to be smaller in TRL group. Nitric oxide synthesis inhibition with L-NAME reduced the baroreflex sensitivity in sedentary and trained animals. Conclusion: Our results showed that nitric oxide synthesis blockade impaired the cardiovascular autonomic adaptations induced by previous aerobic physical training in rats that might be, at least in part, ascribed to a decreased baroreflex sensitivity. (C) 2009 Elsevier B.V. All rights reserved.

Development and characterization of 15 microsatellite loci for Cariniana estrellensis and transferability to Cariniana legalis, two endangered tropical tree species

From a genomic library enriched for GA/CA repeats, 15 highly polymorphic microsatellite markers were developed for Cariniana estrellensis, a tropical forest tree. The microsatellite loci were screened in 49 mature trees found between Pardo river and Mogi-Gua double dagger u river basins, in the state of So Paulo, Brazil. A total of 140 alleles were detected with an average of 9.33 alleles per locus. The expected heterozygosity ranged from 0.37 to 0.88. These loci showed a high probability of paternity exclusion. Additionally, 12 loci were effectively transferred to Cariniana legalis. High levels of polymorphism make the present SSR markers useful for population genetic studies.