Regulation of uteroplacental blood flow (UPBF) during pregnancy remains unclear. Large conductance, Ca(2+)-activated K(+) channels (BK(Ca)), consisting of alpha- and regulatory beta-subunits, are expressed in uterine vascular smooth muscle (UVSM) and contribute to the maintenance of UPBF in the last third of ovine pregnancy, but their expression pattern and activation pathways are unclear. We examined BK(Ca) subunit expression, the cGMP-dependent signaling pathway, and the functional role of BK(Ca) in uterine arteries (UA) from nonpregnant (n = 7), pregnant (n = 38; 56-145 days gestation; term, approximately 150 days), and postpartum (n = 15; 2-56 days) sheep. The alpha-subunit protein switched from 83-87 and 105 kDa forms in nonpregnant UVSM to 100 kDa throughout pregnancy, reversal occurring >30 days postpartum. The 39-kDa beta(1)-subunit was the primary regulatory subunit. Levels of 100-kDa alpha-subunit rose approximately 70% during placentation (P < 0.05) and were unchanged in the last two-thirds of pregnancy; in contrast, beta(1)-protein rose throughout pregnancy (R(2) = 0.996; P < 0.001; n = 13), increasing 50% during placentation and approximately twofold in the remainder of gestation. Although UVSM soluble guanylyl cyclase was unchanged, cGMP and protein kinase G(1alpha) increased (P < 0.02), paralleling the rise and fall in beta(1)-protein during pregnancy and the puerperium. BK(Ca) inhibition not only decreased UA nitric oxide (NO)-induced relaxation but also enhanced alpha-agonist-induced vasoconstriction. UVSM BK(Ca) modify relaxation-contraction responses in the last two-thirds of ovine pregnancy, and this is associated with alterations in alpha-subunit composition, alpha:beta(1)-subunit stoichiometry, and upregulation of the cGMP-dependent pathway, suggesting that BK(Ca) activation via NO-cGMP and beta(1) augmentation may contribute to the regulation of UPBF.

These are the first data demonstrating that BK(Ca) are essential in the maintenance of basal UBF in the last third of ovine pregnancy.

Vascular smooth muscle (VSM) maturation is developmentally regulated and differs between vascular beds. The maturation and contribution of VSM function to tissue blood flow and blood pressure regulation during early gestation are unknown. The carotid artery (CA) contributes to fetal cerebral blood flow regulation and well being. We studied CA VSM contractility, protein contents, and phenotype beginning in the midthird of ovine development. CAs were collected from early (88 -101 day of gestation) and late (138 -150 day; term ϭ day 150) fetal (n ϭ 14), newborn (6 -8 day old; n ϭ 7), and adult (n ϭ 5) sheep to measure forces in endothelium-denuded rings with KCl, phenylephrine, and ANG II; changes in cellular proteins, including total and soluble protein, actin and myosin, myosin heavy chain isoforms (MHC), filamin, and proliferating cell nuclear antigen; and vascular remodeling. KCl and phenylephrine elicited age-and dose-dependent contraction responses (P Ͻ 0.001) at all ages except early fetal, which were unresponsive. In contrast, ANG II elicited dose responses only in adults, with contractility increasing greater than fivefold vs. that shown in fetal or neonatal animals (P Ͻ 0.001). Increased contractility paralleled age-dependent increases (P Ͻ 0.01) in soluble protein, actin and myosin, filamin, adult smooth muscle MHC-2 (SM2) and medial wall thickness and reciprocal decreases (P Ͻ 0.001) in nonmuscle MHC-B, proliferating cell nuclear antigen and medial cellular density. VSM nonreceptor-and receptor-mediated contractions are absent or markedly attenuated in midgestation and increase age dependently, paralleling the transition from synthetic to contractile VSM phenotype and, in the case of ANG II, paralleling the switch to the AT1 receptor. The mechanisms regulating VSM maturation and thus blood pressure and tissue perfusion in early development remain to be determined. myosin heavy chain isoforms; nonmuscle myosin; fetal development; receptor and nonreceptor function; smooth muscle growth; angiotensin II SMOOTH MUSCLE DEVELOPMENT normally proceeds in a wellorchestrated manner before and after birth (5,6,11,14,41,53). These changes occur in three phases: cellular differentiation, functional maturation, and growth (42). During differentiation, progenitor cells derived from the mesenchyme are transformed into immature smooth muscle cells (SMC) localized to either visceral or vascular sites. The subsequent maturational changes result in developmentally regulated improvements in specific organ or vascular function essential for the well being and growth of the developing fetus and newborn. For example, functional maturation of the ovine bladder and umbilical artery smooth muscle occurs early in development (4, 6). The former is required for maintenance of fetal fluid balance and establishment of amniotic fluid volume, which permit normal lung development. The latter is essential in the regulation of fetal oxygen and nutrient uptake from the maternal placental circulation and probably blood pressure (4, 32). In co...

Large conductance K + channels (BK Ca ) are expressed in uterine artery (UA) smooth muscle from nonpregnant and pregnant sheep and contribute to the regulation of basal vascular tone and responses to estrogen and vasoconstrictors. To determine if BK Ca are expressed in women and contribute to UA function, we collected UA from nonpregnant women (n=31) at elective hysterectomy and analyzed for subunit protein, localization with immunohistochemistry and function using endothelium-denuded rings. UA expresses BK Ca α-, β1-and β2-subunit protein. KCl and phenylephrine (PE, an α 1 -agonist) caused dose-dependent vasoconstriction (P<0.001), and UA precontracted with PE dose-dependently relaxed with sodium nitroprusside (SNP; P<0.001). Tetraethylammonium chloride (TEA, 0.2-1.0 mM), a BK Ca inhibitor, dose-dependently increased resting tone (P=0.004; 28±5.3% with 1.0 mM), enhanced PE-induced (10 −6 M) vasoconstriction (P<0.04), and attenuated SNP-induced relaxation at 1.0 mM (P=0.02). BK Ca are expressed in human UA and modulate vascular function by attenuating vasoconstrictor responses and contributing to nitric oxide-induced vasorelaxation.

Background Muscle loss is a sequela of severe burn and critical illness with bed rest contributing significantly to atrophy. We hypothesize that exercise will mitigate muscle loss after burn with bed rest. Materials and Methods Male rats were assigned to sham ambulatory (S/A), burn ambulatory (B/A), sham hindlimb unloading (S/H), or burn hindlimb unloading (B/H). Rats received a 40% scald burn or sham and were ambulatory or placed in hindlimb unloading, a model of bed rest. Half performed twice-daily resistance climbing. Hindlimb isometric forces were measured on day 14. Results Soleus mass and muscle function were not affected by burn alone. Mass was significantly lower in hindlimb unloading (79 vs.139 mg, p<0.001) and no exercise (103 vs.115 mg, p<0.01). Exercise significantly increased soleus mass in B/H (86 vs. 77mg, p<0.01). Hindlimb unloading significantly decreased muscle force in the twitch (31 vs. 12g, p<0.001), tetanic (55 vs. 148 g, p<0.001), and specific tetanic measurements (12 vs. 22 N/cm2, p<0.001). Effects of exercise on force depended on other factors. In B/H, exercise significantly increased twitch (14 vs. 8 g, p<0.05) and specific tetanic force (14 vs. 7 N/cm2, p<0.01). Fatigue index was lower in ambulatory (55%) and exercise (52%) versus hindlimb (69%, p=0.03) and no exercise (73%, p=0.002). Conclusions Hindlimb unloading is a significant factor in muscle atrophy. Exercise increased the soleus muscle mass, twitch, and specific force in this model. However, the fatigue index decreased with exercise in all groups. This suggests exercise contributes to functional muscle change in this model of disuse and critical illness.

Mean arterial pressure (MAP) increases after birth, however, the mechanisms remain unclear. Systemic angiotensin II (ANG II) infusions increase MAP in newborn sheep, but the direct effects of ANG II on peripheral vascular resistance (PVR) are minimal. Thus, its systemic pressor effects may reflect release of other pressor agents, e.g. ␣-agonists and/or AVP, suggesting they contribute to postnatal regulation of MAP and PVR. To address this, we performed studies in conscious sheep at 7-14, 15-21, and 22-35 d postnatal, infusing phenylephrine (PE) or AVP systemically or intra-arterially into the hindlimb while measuring MAP, heart rate (HR), and femoral blood flow (FmBF The transition at birth is accompanied by increases in MAP that occur in at least two phases. There is an initial rapid rise soon after birth, likely due to the removal of the low-resistance placental vascular bed and closure of several fetal vascular shunts. Simultaneous with this is an elevation in circulating levels of catecholamines, ANG II, and AVP (1-4). The second phase is characterized by a gradual rise in MAP that occurs over several days and appears to be developmentally regulated (1,5-9). The mechanisms regulating these changes in MAP are poorly understood and could include the postnatal clearance of placentally derived vasodilators, e.g. prostacyclin, nitric oxide, and estrogen (10,11), increases in central sympathetic outflow (12), age-dependent changes in VSM receptor expression or function (13), maturational and/or function changes in VSM (14), or increases in cardiac output (15). Although cardiac output increases in absolute terms, values (measured in milliliter per minute per kilogram) are reported to fall in the first month after birth (15).Alternatively, changes in either the availability or vascular responsiveness to circulating vasoactive substances may contribute to the regulation of postnatal MAP. For example, ANG II, catecholamines, and AVP modulate MAP in the fetus and adult (1,16 -19) and might also contribute to postnatal regulation of MAP. This could occur through an increase in their direct effects on PVR, which might be due to decreases in the synthesis of local vasodilators (10,11) or increases in VSM sensitivity.Indeed, the responses to norepinephrine and AVP in the isolated ear artery of the developing sheep increase with increasing postnatal age (20

Sensitivity to α-stimulation exceeds ANG II in NP and P UA, explaining the differential uteroplacental sensitivity in pregnancy. Because AT(2)R predominate in UA VSM throughout reproduction, this contributes to the inherent refractoriness to ANG II in the uterine vasculature. The increase in UA contractile proteins at term P suggests remodeling, explaining the enhanced contractility seen.

The role of the renin-angiotensin system (RAS) in regulating newborn mean arterial blood pressure (MAP) and tissue blood flow remains unclear. Although postnatal MAP increases, vascular responsiveness to infused angiotensin II (ANG II) is unchanged, possibly reflecting increased metabolic clearance rate of ANG II (MCR(ANG II)). To address this, we examined MAP, heart rate, plasma ANG II and renin activity (PRA), and MCR(ANG II) in conscious postnatal sheep (n = 9, 5-35 d old) before and during continuous systemic ANG II infusions to measure MCR (ANG II). Postnatal MAP increased (p < 0.02), whereas plasma ANG II decreased from 942 +/- 230 (SEM) to 471 +/- 152 and 240 +/- 70 pg/mL at <10 d, 10-20 d, and 21-35 d postnatally (p = 0.05), respectively. Despite high plasma ANG II, PRA remained elevated, averaging 6.70 +/- 1.1 ng/mL.h throughout the postnatal period, but decreased 35% (p = 0.01) during ANG II infusions. MCR(ANG II) decreased approximately sixfold after birth and averaged 115 mL/min.kg during the first month. Circulating ANG II is markedly increased after birth, reflecting placental removal, high fetal MCR(ANG II), and enhanced RAS activity. Although circulating ANG II decreases as MAP increases, MCR(ANG II) is unchanged, suggesting decreased ANG II production. Persistent vascular smooth muscle (VSM) AT2 receptor subtype (AT2R) expression after birth may modify the hypertensive effects of ANG II postnatally.