We examined the effects of flow on lymphatic endothelial cells by using conventional cascade preparations of isolated coronary arteries without intact endothelium. The pressurized thoracic ducts were intraluminally perfused at a constant flow rate ranging from 0.5 to 2.0 ml/min. A linear relationship was observed between the flow rate and the normalized amount of relaxing substance(s) released from the lymphatic endothelial cells. Thus the flow rate of 2.0 ml/min produced ∼39% of sodium nitroprusside (SNP)–produced maximal relaxation in the cascade arterial rings. The acetylcholine (ACh, 10⁻⁵ M)- and flow-induced relaxations of the cascade arterial rings were completely reduced by the mechanical rubbing of lymphatic endothelial cells in the pressurized lymph vessels. Pretreatment with 5×10⁻⁵ M N^G-nitro-L-arginine methyl ester (L-NAME) on the lymphatic endothelial cells caused a significant reduction of the ACh- and flow-induced vasodilations of the cascade arterial rings. Pretreatment with 10⁻⁵ M indomethacin on the lymphatic endothelial cells produced no significant effect on the ACh- and flow-induced vasodilations. These findings suggest that lymphatic endothelial cells of canine thoracic ducts can produce and release endogenous nitric oxide by stimulation of flow (∼2.0 ml/min).

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Calcium channels are essential for excitation-contraction coupling and pacemaker potentials in cardiac muscle cells. Whereas L-type Ca²⁺ channels have been extensively studied, T-type channels have been poorly characterized in cardiac myocytes. We describe here the functional properties of recombinant Ca_V3.2 T-type Ca²⁺ channels expressed in mammalian cell lines. The T-type Ca²⁺ current showed a rapid activation and an inactivation phase in response to depolarization, and it displayed a window current over the voltage range from −60 to −40 mV in 1 to 10 mM external Ca²⁺. Barium (Ba²⁺) and strontium (Sr²⁺) permeate the channel with similar activation kinetics. On the other hand, monovalent cations, Li⁺ and Na⁺, permeate the T-type Ca²⁺ channel more easily than the L-type Ca²⁺ channel. The permeability order of the Ca_V3.2 T-type Ca²⁺ channel among monovalent and divalent cations was determined as Ba²⁺>Mn²⁺>Ca²⁺>Sr²⁺>Li⁺¹>Na⁺ with the permeability order of 1.39:1.25:1.00:0.95:0.55:0.29. The ionic conductance sequence for cations relative to calcium was Sr²⁺>Ba²⁺>Ca²⁺>Li⁺¹>Mn²⁺>Na⁺ with the conductance ratio of 1.39:1.21:1.00:0.40:0.23:0.11. The permeation profile of manganese (Mn²⁺) is complex. Mn²⁺ permeates the Ca²⁺ channel with a permeability similar to Ca²⁺ or Ba²⁺, but with a much smaller current density, resulting in a much smaller conductance. The properties relating to progression and recovery from inactivation in the Ca_V3.2 channel are substantially identical with either Ca²⁺ or Ba²⁺ as the charge carrier.

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The objective of the present study was to investigate the differential activation of protein kinase C between ischemic (IPC) and pharmacological preconditioning (PPC) in the rabbit heart. Control, IPC, diazoxide (Diaz), and chelerythrine (Chel)+IPC groups underwent prolonged coronary artery occlusion (CAO) for 30 minutes followed by 180 minutes' reperfusion (protocol I). In protocol II, sham, IPC-only, Diaz-only, and Chel+IPC-only groups did not undergo prolonged CAO. IPC was induced with 4 cycles of 5-min regional ischemia and 10-min reperfusion before prolonged CAO. Diaz (5 mg/kg) was administered 30 min before prolonged CAO. Chel (5 mg/kg) was administered 5 min before the IPC procedure. Infarct size was determined by tetrazolium staining. Assessment of protein kinase C (PKC) isoforms from a left ventricular (LV) sample was conducted by western blotting. Apoptosis in situ was determined by TUNEL assay. The infarction area in the IPC (11.6 ± 1.0%) and Diaz (19.5 ± 3.8%) groups was reduced significantly (plt;0.01, plt;0.05) relative to the control group (40.0 ± 3.8%). The reduction by IPC was abolished by pretreatment with Chel. Apoptosis was significantly decreased (plt;0.01) in the IPC and diazoxide groups compared with the control and Chel+IPC groups (control: 4.78 ± 0.56% vs. IPC: 2.00 ± 0.38% vs. Diaz: 2.20 ± 0.32% vs. Chel+IPC: 4.32 ± 0.41%) and DNA laddering was attenuated in the IPC and Diaz groups. Membrane PKC-ε levels in the IPC and Diaz groups increased significantly relative to the control and Chel+IPC groups. Membrane PKC-ε levels in the IPC-only group showed greater increases than the Diaz-only and Chel+IPC-only groups. These findings suggest that whereas PPC suppresses apoptosis when diazoxide opens mitochondrial K_ATP channels and then activates PKC-ε through ischemia-reperfusion, IPC activates PKC-ε in the particulate fraction prior to continuous ischemia-reperfusion. We concluded that the difference between IPC and PPC appears to consist in the difference in the timing of PKC-ε activation, though both IPC and PPC provide the cardioprotection in ischemia-reperfusion injury.

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Capillary proliferation occurs during compensatory hypertrophy. We investigated whether the expression of vascular endothelial growth factor (VEGF) is elevated at the onset of hypertrophy when capillary proliferation is minimal, and whether muscle damage as assessed by muscle force deficits, may occur at the onset of hypertrophy. To investigate this, we induced in 9-month-old rats, under isoflurane anesthesia, hypertrophy of the left plantaris muscle by denervation of the gastrocnemius and soleus muscles. Capillarization was investigated in both the deep (oxidative) and the superficial (glycolytic) regions of the plantaris muscle. After 2 weeks, muscle mass had increased by 16% (plt;0.01), which was not accompanied by increases in fiber size. The maximal tetanic force (P₀) and specific tension (P₀·g⁻¹ or P₀·cm²) and twitch characteristics were unaltered, and fatigue resistance of the overloaded muscle was improved (plt;0.05). However, the myosin heavy chain composition was unaltered. Capillary proliferation was not yet evident, but VEGF mRNA and protein levels were elevated 1.5- and 8-fold, respectively (plt;0.05). We concluded that the normal specific tension and the elevated VEGF expression after 2 weeks of overload indicate (1) an absence of or minimal muscle damage at this early time point, and (2) that elevated VEGF expression precedes and is involved in capillary proliferation that occurs during the later stages of compensatory hypertrophy.

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We examined chronic effects of 17β-estradiol (E₂β) on the responses of isolated rat anterior cerebral small arteries to vasoactive substances with special reference to endothelial function. Female Sprague-Dawley rats were separated into four groups: (1) sham-operated group (Sham), (2) sham-operated plus E₂β treated group (Sham+E), (3) ovariectomized group (OVX), (4) ovariectomized plus E₂β treated group (OVX+E). 5-Hydroxytryptamine (5-HT) (10⁻¹⁰–10⁻³ M) and U46619 (10⁻¹⁵–10⁻⁸ M) induced concentration-dependent contractions in the cerebral small arteries. The 5-HT- and U46619-induced contractions were not affected by pretreatment with 3×10⁻⁵ M N^ω-nitro-L-arginine methyl ester (L-NAME). No significant difference in high potassium (80 mM)- and the agonists-mediated contractions was observed among the four groups. Administration of acetylcholine (ACh) (10⁻⁹–10⁻³ M) and sodium nitroprusside (SNP) (10⁻⁸–10⁻³ M) caused dose-related relaxations in the cerebral small arteries precontracted by 10⁻⁸ M U46619. Chronic treatment with E₂β caused a significant potentiation of the ACh-induced relaxations in the Sham+E and OVX+E groups. The dose–response curve for ACh in the OVX group was quite similar to that obtained with the Sham group. The ACh-induced relaxation was reduced significantly by pretreatment with 3×10⁻⁵ M L-NAME, and an additional treatment with 10⁻³ M L-arginine reversed significantly the L-NAME–induced inhibition. The removal of endothelial cells produced a significant reduction of the ACh-induced relaxation. Indomethacin (10⁻⁵ M) did not alter the ACh-induced relaxation. The findings suggest that E₂β potentiates ACh-induced endothelium-dependent relaxation in rat anterior cerebral arteries and that the potentiation may be, in part, mediated by increasing production and release of endogenous NO from the endothelial cells.

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Repeat immobilization-stressed rats are leaner and have improved cold tolerance due to enhancement of brown adipose tissue (BAT) thermogenesis. This process likely involves stress-induced sympathetic nervous system activation and adrenocortical hormone release, which dynamically enhances and suppresses uncoupling protein 1 (UCP1) function, respectively. To investigate whether repeated immobilization influences UCP1 thermogenic properties, we assessed UCP1 mRNA, protein expression, and activity (GDP binding) in BAT from immobilization-naive or repeatedly immobilized rats (3 h daily for 4 weeks) and sham operated or adrenalectomized (ADX) rats. UCP1 properties were assessed before (basal) and after exposure to 3 h of acute immobilization. Basal levels of GDP binding and UCP1 expression was significantly increased (140 and 140%) in the repeated immobilized group. Acute immobilization increased GDP binding in both naive (180%) and repeated immobilized groups (220%) without changing UCP1 expression. In ADX rats, basal GDP binding and UCP1 gene expression significantly increased (140 and 110%), and acute immobilization induced further increase. These data demonstrate that repeated immobilization resulted in enhanced UCP1 function, suggesting that enhanced BAT thermogenesis contributes to lower body weight gain through excess energy loss and an improved ability to maintain body temperature during cold exposure.

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Since human thermoregulatory heat loss responses, cutaneous vasodilation and sweating, cause hypovolemia, they should resultantly stimulate human baroreflexes. However, it is possible that the thermoregulatory system directly interacts with the baroreflex system through central neural connections independently of the heat-induced hypovolemia. We hypothesized that heat stress modifies the baroreflex control of sympathetic nerve activity independently of heat-induced hypovolemia in humans. We made whole-body heating with tube-lined suits perfused with warm water (46–47°C) on 10 healthy male subjects. The heating increased skin and tympanic temperatures by 10.0 and 0.4°C, respectively. It increased resting total muscle sympathetic nerve activity (MSNA, microneurography) by 94 ± 9% and decreased central venous pressure (CVP, dependent arm technique) by 2.6 ± 0.9 mmHg. The heating increased arterial baroreflex gain by 193%, assessed as a response of MSNA to a decrease in diastolic arterial pressure during Valsalva's maneuver, but it did not change threshold arterial pressure for MSNA activation. Although the heating did not change the cardiopulmonary baroreflex gain assessed as a response of MSNA to a change in estimated central venous pressure (CVP) during a 10° head-down and -up tilt test, it upwardly shifted the stimulus-response baroreflex relationship. These changes in baroreflex functions during heating were not restored by an intravenous infusion of warmed isotonic saline (37°C, 15 ml/kg) that restored the heat-induced reduction of CVP. Our results support our hypothesis that heat stress modifies the baroreflex control of MSNA independently of heat-induced hypovolemia in humans. Our results also suggest that the hyperthermal modification of baroreflex results from central neural interaction between thermoregulatory and baroreflex systems. [Japanese Journal of Physiology, 53, 215–222, 2003]

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To evaluate changes in the cerebral circulation during acute microgravity (μG), we measured intracranial pressure (ICP), aortic pressure at the diaphragm level, and cerebral flow velocity (CFV) in anesthetized rats (n = 5) during 4.5 s of μG induced by free drop, then calculated arterial pressure at the eye level (AP_eye) and cerebral perfusion pressure (CPP = AP_eye−ICP), and estimated CPP–CFV relationship. The rats were placed in the flat and the 30° head-up positions. In the head-up position, ICP, AP_eye, and CPP were significantly increased by 2.2 ± 0.4, 12.3 ± 2.0, and 10.1 ± 1.7 mmHg respectively during μG, whereas the CFV did not change significantly. In the flat position, none of these variables were significantly affected by μG. The slope of the CPP–CFV relationship was decreased only in the head-up position, suggesting that the cerebrovascular resistance was increased by μG. These findings indicate that the change in gravitational (hydrostatic) pressure is a key factor in understanding the changes in cerebral circulation during acute μG.

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The effects of beta adrenergic agonists, clenbuterol (2 mg/kg body weight/d) and isoproterenol (12 mg/kg body weight/d), in normal innervated and denervated rat gastrocnemius muscle were investigated. The daily administration of beta adrenergic agonists to normal innervated rats for a short period (7 d) resulted in the hypertrophy of gastrocnemius as confirmed from the measurement of total tissue protein contents. The development of denervation atrophy witnessed a stimulation in the expression of acid and alkaline phosphatases, pointing to an enhanced myofibrillar degeneration. An administration of beta adrenergic agonists inhibited the expression of raised levels of these enzymes in denervated muscle. A measurement of 3-methylhistidine in muscle revealed a loss of amino acid with the progress in the development of denervation atrophy. Serum and urine samples from denervated rats showed a progressive accumulation of 3-methylhistidine. Clenbuterol and isoproterenol treatment to these rats resulted in an inhibition of 3-methylhistidine accumulation. When 3-methylhistidine was used as a marker of myofibrillar degeneration, the results seemed to suggest that the degeneration of cyto-contractile apparatus accompanying denervation atrophy is attenuated in the presence of beta adrenergic agonists, implying that these sympathomimetic drugs are capable of reversing denervation atrophy in rat gastrocnemius.

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We examined the effect of low-intensity single-leg exercise (20 or 30 watt, 5 min) on pulse wave velocity from the femoral to the ankle arteries in 18 young men. After the exercise, the velocity significantly decreased in the exercised leg, but not in the counterpart, suggesting that the decrease in arterial stiffness in the exercised leg was induced mainly by exercise-related regional factors.

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Rib cage inspiratory muscle contractions at a deep inflated position caused a subsequent increase in rib cage cross-sectional area at the end-expiratory phase. The increase in this area was enhanced by the activation of muscle spindles in the inspiratory muscles by a mechanical vibration combined with muscle contractions.

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Pathophysiological roles of apoptosis in post-infarction left ventricular (LV) remodeling have not been well characterized. This study showed that TUNEL- or cleaved caspase-3–positive myocytes were identified late after ligation of the left coronary artery in rats, suggesting that apoptotic myocyte death contributed to the morphological change associated with LV remodeling.

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