Sergio Rey

Chronic intermittent hypoxia enhances cat chemosensory and ventilatory responses to hypoxia

The carotid body (CB) chemoreceptors may play an important role in the enhanced hypoxic ventilatory response induced by chronic intermittent hypoxia (CIH). We studied the effects of cyclic hypoxic episodes of short duration on cat cardiorespiratory reflexes, heart rate variability, and CB chemosensory activity. Cats were exposed to cyclic hypoxic episodes (P  O 2 ∼ 75 Torr) repeated during 8 h for 2–4 days. Cats were anaesthetized with sodium pentobarbitone (40 mg kg−1i.p., followed by 8–12 mg i.v.), and ventilatory and cardiovascular responses to NaCN (0.1–100 μg kg−1i.v.) and several isocapnic levels of oxygen (P  O 2 ∼ 20–740 Torr) were studied. After studying the reflex responses, we recorded the CB chemosensory responses induced by the same stimuli. Results showed that CIH for 4 days selectively enhanced cat CB ventilatory (VT and VI) responses to hypoxia, while responses to NaCN remained largely unchanged. Similarly, basal CB discharges and responses to acute hypoxia (P  O 2 < 100 Torr) were larger in CIH than in control cats, without modification of the responses to NaCN. Exposure to CIH did not increase basal arterial pressure, heart rate, or their changes induced by acute hypoxia or hyperoxia. However, the spectral analysis of heart rate variability of CIH cats showed a marked increase of the low‐/high‐frequency ratio and an increase of the power spectral distribution of low frequencies of heart rate variability. Thus, the enhanced CB reactivity to hypoxia may contribute to the augmented ventilatory response to hypoxia, as well as to modified heart rate variability due to early changes in autonomic activity.

Contribution of endothelin-1 to the enhanced carotid body chemosensory responses induced by chronic intermittent hypoxia.

Chronic intermittent hypoxia (CIH) enhances carotid body (CB) chemosensory responses to acute hypoxia. We tested the hypothesis that endothelin-1 (ET-1), an excitatory modulator of CB chemoreception may contribute to the enhanced CB chemosensory responses in cats exposed to cyclic hypoxic episodes repeated during 8 h for 4 days. Accordingly, we measured the ET-1 immunoreactivity (ET-ir) in the CB and plasma. Using a perfused CB preparation, we studied the effects of exogenous ET-1 and bosentan, a non-selective endothelin receptor type A and B antagonist, on the frequency of chemosensory discharges (f(x)) during normoxia, mild and severe hypoxia. We found that CIH increased ET-ir in the CB by approximately 10-fold leaving ET-1 plasma levels unchanged. Application of ET-1 to control and CIH-treated CBs produced long-lasting dose-dependent increases in f(x), although the dose-response curve showed a rightward-shift in the CIH-treated CBs. CIH increased baseline f(x) and hypoxic chemosensory responses, which were reduced by 50 microM bosentan in CBs from CIH-treated cats. Present results suggest that a local increase of ET-1 in the CB may contribute to the enhanced chemosensory responses induced by CIH predominantly through a vasomotor mechanism.

Lipopolysaccharide‐induced carotid body inflammation in cats: functional manifestations, histopathology and involvement of tumour necrosis factor‐α

In the absence of information on functional manifestations of carotid body (CB) inflammation, we studied an experimental model in which lipopolysaccharide (LPS) administration to pentobarbitone‐anaesthetized cats was performed by topical application upon the CB surface or by intravenous infusion (endotoxaemia). The latter caused: (i) disorganization of CB glomoids, increased connective tissue, and rapid recruitment of polymorphonuclear cells into the vascular bed and parenchyma within 4 h; (ii) increased respiratory frequency and diminished ventilatory chemoreflex responses to brief hypoxia (breathing 100% N2 for 10 s) and diminished ventilatory chemosensory drive (assessed by 100% O2 tests) during normoxia and hypoxia; (iii) tachycardia, increased haematocrit and systemic hypotension in response to LPS i.v.; and (iv) increased basal frequency of carotid chemosensory discharges during normoxia, but no change in maximal chemoreceptor responses to brief hypoxic exposures. Lipopolysaccharide‐induced tachypnoea was prevented by prior bilateral carotid neurotomy. Apoptosis was not observed in CBs from cats subjected to endotoxaemia. Searching for pro‐inflammatory mediators, tumour necrosis factor‐α (TNF‐α) was localized by immunohistochemistry in glomus and endothelial cells; reverse transcriptase‐polymerase chain reaction revealed that the CB expresses the mRNAs for both type‐1 (TNF‐R1) and type‐2 TNF‐α receptors (TNF‐R2); Western blot confirmed a band of the size expected for TNF‐R1; and histochemistry showed the presence of TNF‐R1 in glomus cells and of TNF‐R2 in endothelial cells. Experiments in vitro showed that the frequency of carotid nerve discharges recorded from CBs perfused and superfused under normoxic conditions was not significantly modified by TNF‐α, but that the enhanced frequency of chemosensory discharges recorded along responses to hypoxic stimulation was transiently diminished in a dose‐dependent manner by TNF‐α injections. The results suggest that the CB may operate as a sensor for immune signals, that the CB exhibits histological features of acute inflammation induced by LPS, that TNF‐α may participate in LPS‐induced changes in chemosensory activity and that some pathophysiological reactions to high levels of LPS in the bloodstream may originate from changes in CB function.

Cardiovascular and ventilatory acclimatization induced by chronic intermittent hypoxia: A role for the carotid body in the pathophysiology of sleep apnea

Patients with obstructive sleep apnea (OSA) show augmented ventilatory, sympathetic and cardiovascular responses to hypoxia. The facilitatory effect of chronic intermittent hypoxia (CIH) on the hypoxic ventilatory response has been attributed to a potentiation of the carotid body (CB) chemosensory response to hypoxia. However, it is a matter of debate whether the effects induced by CIH on ventilatory responses to hypoxia are due to an enhanced CB activity. Recently, we studied the effects of short cyclic hypoxic episodes on cat cardiorespiratory reflexes, heart rate variability, and CB chemosensory activity. Cats were exposed to cyclic hypoxic episodes repeated during 8 hours for 4 days. Our results showed that CIH selectively enhanced ventilatory and carotid chemosensory responses to acute hypoxia. Exposure to CIH did not increase basal arterial pressure, heart rate, or their changes induced by acute hypoxia. However, the spectral analysis of heart rate variability of CIH cats showed a marked increase of the low/high frequency ratio and an increased variability in the low frequency band of heart rate variability, similar to what is observed in OSA patients. Thus, it is likely that the enhanced CB reactivity to hypoxia may contribute to the augmented ventilatory response to hypoxia.

Expression and immunolocalization of endothelin peptides and its receptors, ETA and ETB, in the carotid body exposed to chronic intermittent hypoxia.

Increased levels of endothelin-1 (ET-1) in the carotid body (CB) contribute to the enhancement of chemosensory responses to acute hypoxia in cats exposed to chronic intermittent hypoxia (CIH). However, it is not known if the ET receptor types A (ETA-R) and B (ETB-R) are upregulated. Thus, we studied the expression and localization of ETA-R and ETB-R using Western blot and immunohistochemistry (IHC) in CBs from cats exposed to cyclic hypoxic episodes, repeated during 8 hr for 4 days. In addition, we determined if ET-1 is expressed in the chemoreceptor cells using double immunofluorescence for ET-1 and tyrosine hydroxylase (TH). We found that ET-1 expression was ubiquitous in the blood vessels and CB parenchyma, although double ET-1 and TH-positive chemoreceptor cells were mostly found in the parenchyma. ETA-R was expressed in most chemoreceptor cells and blood vessels of the CB vascular pole. ETB-R was expressed in chemoreceptor cells, parenchymal capillaries, and blood vessels of the vascular pole. CIH upregulated ETB-R expression by ∼2.1 (Western blot) and 1.6-fold (IHC) but did not change ETA-R expression. Present results suggest that ET-1, ETA-R, and ETB-R are involved in the enhanced CB chemosensory responses to acute hypoxia induced by CIH.

Dynamic time-varying analysis of heart rate and blood pressure variability in cats exposed to short-term chronic intermittent hypoxia

Chronic intermittent hypoxia (CIH) contributes to the development of hypertension in patients with obstructive sleep apnea and animal models. However, the early cardiovascular changes that precede CIH-induced hypertension are not completely understood. Nevertheless, it has been proposed that one of the possible contributing mechanisms to CIH-induced hypertension is a potentiation of carotid body (CB) hypoxic chemoreflexes. Therefore, we studied the dynamic responses of heart rate, blood pressure, and their variabilities during acute exposure to different levels of hypoxia after CIH short-term preconditioning (4 days) in cats. In addition, we measured baroreflex sensitivity (BRS) on the control of heart rate by noninvasive techniques. To assess the relationships among these indexes and CB chemoreflexes, we also recorded CB chemosensory discharges. Our data show that short-term CIH reduced BRS, potentiated the increase in heart rate induced by acute hypoxia, and was associated with a dynamic shift of heart rate variability (HRV) spectral indexes toward the low-frequency band. In addition, we found a striking linear correlation (r = 0.97) between the low-to-high frequency ratio of HRV and baseline. CB chemosensory discharges in the CIH-treated cats. Thus, our results suggest that cyclic hypoxic stimulation of the CB by short-term CIH induces subtle but clear selective alterations of HRV and BRS in normotensive cats.

Expression of kallikrein, bradykinin B2 receptor, and endothelial nitric oxide synthase in placenta in normal gestation, preeclampsia, and placenta accreta

In an effort to define the varied expression of three vasoactive markers in the clinical models of normal placenta/normal invasion (n=11), preeclampsia/restricted trophoblast invasion (n=15), and placenta accreta/exaggerated invasion (n=6), we performed semiquantitative immunohistochemistry for kallikrein, bradykinin B2 receptor, and endothelial nitric oxide synthase (eNOS). In the floating villi, the syncytiotrophoblast expressed more kallikrein in placenta accreta (p<0.05), than in normal and preeclamptic placentas, while the bradykinin B2 receptor and eNOS were similarly expressed in all groups; in the fetal endothelium, the bradykinin B2 receptor was enhanced in placenta accreta (p<0.005), but kallikrein and eNOS were similarly expressed in the other two groups. In the extravillous trophoblast, both kallikrein and eNOS expression were higher in placenta accreta (p<0.001), while the bradykinin B2 receptor signal was only enhanced in preeclampsia (p<0.05). The presence and localization of kallikrein, the bradykinin B2 receptor, and eNOS in the fetomaternal interface in the three study conditions supports a local role for interrelated vasodilatory/antiaggregating systems. This first report of the variations observed in kallikrein and eNOS in a condition of exaggerated trophoblast invasion supports the participation of vasodilatation in trophoblast migration.

Endothelins and Nitric Oxide: Vasoactive Modulators of Carotid Body Chemoreception

The carotid body (CB) is the main arterial chemoreceptor that senses arterial PO2, PCO2 and pH. The structural unit of the CB is the glomoid, which is formed by clusters of chemoreceptor (glomus) cells located around the capillaries. The glomus cells are synaptically connected to nerve terminals of petrosal ganglion (PG) neurons and surrounded by sustentacular cells. The most accepted model of CB chemoreception states that glomus cells are the primary sensors. In response to hypoxia, hypercapnia and acidosis, glomus cells release one or more transmitters, which acting on the nerve terminals of sensory PG neurons, increase the chemosensory discharge. The CB has a high blood flow and an elevated metabolism that correlate to its oxygen-sensing function. Thus, vasoactive molecules produced within the CB may modulate the chemosensory process by controlling the CB blood flow and tissue PO2. In this review, we examine recent evidence supporting the idea that endothelins (ETs) and nitric oxide (NO) modulate the CB function acting upon chemoreceptor cells and chemosensory neurons or by regulating the blood flow through the CB parenchyma.

Spatio-temporal expression of MMP-2, MMP-9 and tissue kallikrein in uteroplacental units of the pregnant guinea-pig (Cavia porcellus)

BackgroundIn humans trophoblast invasion and vascular remodeling are critical to determine the fate of pregnancy. Since guinea-pigs share with women an extensive migration of the trophoblasts through the decidua and uterine arteries, and a haemomonochorial placenta, this species was used to evaluate the spatio-temporal expression of three enzymes that have been associated to trophoblast invasion, MMP-2, MMP-9 and tissue kallikrein (K1).MethodsUteroplacental units were collected from early to term pregnancy. MMP-2, MMP-9 and K1 were analysed by immunohistochemistry and Western blot. The activities of MMP-2 and MMP-9 were assessed by gelatin zymography.ResultsImmunoreactive MMP-2, MMP-9 and K1 were detected in the subplacenta, interlobar and labyrinthine placenta, syncytial sprouts and syncytial streamers throughout pregnancy. In late pregnancy, perivascular or intramural trophoblasts expressed the three enzymes. The intensity of the signal in syncytial streamers was increased in mid and late pregnancy for MMP-2, decreased in late pregnancy for MMP-9, and remained stable for K1. Western blots of placental homogenates at days 20, 40 and 60 of pregnancy identified bands with the molecular weights of MMP-2, MMP-9 and K1. MMP-2 expression remained constant throughout gestation. In contrast, MMP-9 and K1 attained their highest expression during midgestation. Placental homogenates of 20, 40 and 60 days yielded bands of gelatinase activity that were compatible with MMP-2 and MMP-9 activities. ProMMP-2 and MMP-9 activities did not vary along pregnancy, while MMP-2 and MMP-9 increased at 40 and 40–60 days respectively.ConclusionThe spatio-temporal expression of MMPs and K1 supports a relevant role of these proteins in trophoblast invasion, vascular remodeling and placental angiogenesis, and suggests a functional association between K1 and MMP-9 activation.

Endothelins in the cat petrosal ganglion and carotid body: Effects and immunolocalization

In response to hypoxia, chemoreceptor cells of the carotid body (CB) release transmitters, which acting on the petrosal ganglion (PG) neuron terminals, increase the chemoafferent discharge. Additionally, vasoactive molecules produced within the CB may modulate hypoxic chemoreception by controlling blood flow and tissue PO2. Endothelin-1 (ET-1) increases basal CB chemosensory discharges in situ, probably due to its vasoconstrictor action. However, the actions of ET-1 on PG neurons or its expression in the PG are not known. Using immunohistochemistry, we found that endothelin-like peptides are expressed in the cat PG and CB under normoxic conditions. Exogenous applications of ET-1 increased the chemosensory activity in the vascularly perfused CB but were ineffective on either the CB or PG superfused preparations, both of which are devoid of vascular control. Thus, our data indicate that the excitatory effect of ET-1 in the carotid chemoreceptor system appears to be mainly due to a vasoconstrictor effect in the CB blood vessels.