Kohei Kawakami

EFFECTS OF PHYTONCIDES ON BLOOD PRESSURE UNDER RESTRAINT STRESS IN SHRSP

1. Phytoncides are volatile substances released mainly from trees. We studied whether phytoncides can reduce stress responses in stroke‐prone spontaneously hypertensive rats (SHRSP).

Exaggerated response to restraint stress in rats congenic for the chromosome 1 blood pressure quantitative trait locus

1. To understand the roles of a putative hypertension gene in the chromosome 1 quantitative trait locus (QTL) region, the response to restraint stress was studied in strains congenic for this QTL.

Exaggerated response to cold stress in a congenic strain for the quantitative trait locus for blood pressure.

Objective Stroke-prone spontaneously hypertensive rats (SHRSP) are known to have sympathetic hyperactivity to various stimuli. In the search for ‘intermediate phenotypes’ inferring the function of hypertension genes, the present study assessed responsiveness to cold stress in a congenic strain derived from SHRSP/Izm and Wistar–Kyoto/Izm (WKY/Izm). Design A congenic strain, WKYpch1.0, was established by 10 generations of backcrossing to transfer the chromosomal fragment between D1Wox29 and D1Arb21 of SHRSP to WKY. This fragment covered the 100:1 confidence interval of the quantitative trait locus (QTL) for blood pressure identified in a previous study. Response to cold stress was studied by exposing rats to 4°C for 4 h. Blood pressure was monitored with telemetry. Urine was collected during the exposure, and urinary concentrations of catecholamines were measured by high-performance liquid chromatography. Results Under the cold stress, urinary excretion of norepinephrine (NE) and vanillylmandelic acid (VMA), as well as the plasma level of NE, was significantly greater in WKYpch1.0 than in WKY. The increase in blood pressure during the cold stress was also greater in WKYpch1.0 than in WKY. Further, neonatal chemical sympathectomy using guanethidine abolished the exaggerated response in blood pressure and in urinary excretion of NE and VMA in WKYpch1.0. Conclusion These results suggested that the QTL region on rat chromosome 1 harbored genes responsible for the exaggerated response of the sympathetic nervous system to the cold stress. The relationship of this with the pathogenesis of hypertension should be elucidated in future studies.

A 1.8-Mbp fragment on chromosome 1 affects sympathetic response to stress: evaluation in reciprocal congenic strains between stroke-prone spontaneously hypertensive rat and Wistar-Kyoto rat.

Background In the previous studies, we indicated that a gene (or genes) responsible for exaggerated sympathetic response to stress was located in a chromosome 1 QTL for blood pressure (BP) in stroke-prone spontaneously hypertensive rat (SHRSP). In this study, we narrowed down the candidate region to a 1.8-Mbp fragment between D1Rat171 and D1Wox33, and established reciprocal congenic strains for this region. Methods Reciprocal congenic strains were established by introgressing the chromosomal segment from SHRSP/Izm into WKY/Izm (Wpch1.21) and vice versa (SPwch1.72). The urinary norepinephrine excretion (u-NE) was quantified with high-performance liquid chromatography in the urine collected under 6 h of cold stress (4°C). ECG was recorded using the telemetry under 3 h of restraint stress, and the relative sympathetic activity was evaluated as the low frequency/high frequency ratio by the power spectral analysis. BP under the stresses was evaluated by the telemetry. Results The increases in the u-NE during the cold stress and in the low frequency/high frequency ratio under the restraint stress were significantly greater in Wpch1.21 when compared with Wistar–Kyoto (WKY) rat. The increases in BP both under the cold and the restraint stresses were significantly greater in Wpch1.21 than in WKY. In the reciprocal congenic strain, SPwch1.72, the effects of the transferred fragment on the sympathetic stress responses were confirmed as lower u-NE and low frequency/high frequency in this strain than in SHRSP. Further, the BP responses both to the cold and the restraint stresses were significantly greater in SHRSP than in SPwch1.72. Conclusion These results indicated that a small fragment on chromosome 1 harbored a gene (or genes) influencing the sympathetic response to different stresses.

EFFECTS OF DWELLING CONDITIONS ON CARDIOVASCULAR PARAMETERS IN SPONTANEOUSLY HYPERTENSIVE RATS

1 Living conditions are an important environmental factor for laboratory animals. We studied the effects of different living conditions on the cardiovascular parameters in spontaneously hypertensive rats (SHR). 2 In both the light and the dark phase, the heart rate of rats kept in a wire‐mesh cage was significantly greater than the heart rate of rats kept under the other living conditions (with wood‐shaving bedding material in a plastic cage, with cloth bedding material in a plastic cage and with cloth bedding material in a wire‐mesh cage, P < 0.01 by the paired t‐test). 3 Systolic blood pressure and diastolic blood pressure of rats kept in a wire‐mesh cage in the light phase was significantly higher than the heart rates of rats kept under the other living conditions (with wood‐shaving bedding materials in a plastic cage, with cloth bedding materials in a plastic cage, P < 0.01 by the paired t‐test). 4 Based on these results, we concluded that the living conditions had a substantial influence on the cardiovascular parameters in SHR.

Increased Amplitude of the Circadian Variations in Locomotor Activity, Systolic Arterial Pressure, and Heart Rate in Congenic Rats derived from SHRSP Rats

The circadian variations in the hemodynamics and locomotor activity (ACT) of congenic rats derived from stroke-prone spontaneously hypertensive (SHRSP) rats and Wistar-Kyoto (WKY) rats have not been studied in detail. We used radio telemetry and the maximum entropy method to examine these variations. The systolic arterial pressure of the congenic rats was intermediate between those of the SHRSP rats and WKY rats, while their heart rate was lower than that of the SHRSP rats. The congenic rats also showed the highest ACT. The circadian variations in the heart rates of the congenic rats were more like those of the WKY rats, and the variations in their ACT were more similar to those of the SHRSP rats.

A potential contribution of tenascin-X to blood vessel formation in peripheral nerves

Tenascin-X (TNX), an extracellular matrix protein, is abundantly expressed in peripheral nerves. However, the physiological role of TNX in peripheral nerves remains unknown. In this study, we found that actin levels in sciatic nerves of TNX-deficient mice were markedly decreased. Since actin was highly expressed in endothelial cells in wild-type sciatic nerves, we assessed morphological alterations of blood vessels in TNX-null sciatic nerves. The density of blood vessels was significantly decreased and the size of blood vessels was larger than those in wild-type sciatic nerves. Immunofluorescence showed that TNX was expressed by Schwann cells and fibroblasts in sciatic nerves. The results suggest that TNX secreted from Schwann cells and/or fibroblasts is involved in blood vessel formation in peripheral nerves.

Suppression of hepatic dysfunction in tenascin‑X‑deficient mice fed a high‑fat diet

Extracellular matrix glycoprotein tenascin-X (TNX) is the largest member of the tenascin family. In the present study, the contribution of TNX to liver dysfunction was investigated by administration of high-fat and high-cholesterol diet with high levels of phosphorus and calcium (HFCD) to wild-type (WT) and TNX-knockout (KO) mice. After 16 weeks of HFCD administration, the ratio of liver weight to body weight was approximately 22% higher in the HFCD-fed WT mice compared with the HFCD-fed TNX-KO mice, indicating hepatomegaly in HFCD-fed WT mice. Histological analyses with hematoxylin and eosin staining at 21 weeks revealed that hepatocyte hypertrophy in HFCD-fed TNX-KO mice was suppressed to 85% of that in HFCD-fed WT mice. By contrast, there was a 1.2-fold increase in lipid deposition in hepatocytes from HFCD-fed TNX-KO mice compared with HFCD-fed WT mice at 18 weeks, as demonstrated by Oil Red O staining. In addition, TNX-KO mice at 21 weeks and 27 weeks post-HFCD administration exhibited significant suppression of inflammatory cell infiltrate to 51 and 24% of that in WT mice, respectively. Immunofluorescence analysis for type I collagen and Elastica van Gieson staining demonstrated a clear hepatic fibrosis progression in HFCD-fed WT mice at 27 weeks, whereas hepatic fibrosis was undetected in HFCD-fed TNX-KO mice. The present findings indicated that TNX deficiency suppressed hepatic dysfunction induced by HFCD administration.

MPS 13-05 LOCOMOTOR ACTIVITY IN CONGENIC SHRSPwch1.0 RATS IS DERIVED FROM THE INCREASE IN SYSTOLIC ARTERIAL PRESSURE BUT NOT THE SYSTOLIC ARTERIAL PRESSURE LEVEL ITSELF

Objective: Congenic rats (SHRSPwch1.0) were produced from stroke-prone SHR/Izumo rats (SHRSP/Izm) and Wistar-Kyoto rat/Izumo rats (WKY/Izm). We previously found that the systolic arterial pressure (SAP) and heart rate (HR) of the SHRSPwch1.0 were lower than those of the SHRSP/Izm. However, the SHRSPwch1.0 exhibited greater locomotor activity (ACT) than the SHRSP/Izm. The purpose of this study was to investigate the ACT characteristics of SHRSPwch1.0 in a time-series analysis. Design and Method: We used ten mature male SHRSPwch1.0 and ten male age-matched SHRSP/Izm as controls. They were kept under a 12-hour light and dark cycle in identical conditions. The rats’ SAP, HR, and ACT were monitored using radio telemetry, and their ACT was analyzed using the maximum entropy method to enable a spectral analysis of ACT. Results: All of the examined ACT values (24-hour, dark period, and light period) were greater in the SHRSPwch1.0 than in the SHRSP/Izm. A time-series analysis of ACT in the SHRSPwch1.0 revealed both circadian (23.379 hours) and ultradian variations (16.197 hours and 12.184 hours). On the other hand, the ACT of the SHRSP/Izm exhibited circadian variations (23.190 hours) and an almost imperceptible ultradian rhythm. The power spectral density of the ACT spectrum was greater in the SHRSPwch1.0 than in the SHRSP/Izm. A stronger correlation was noted between ACT and SAP in the SHRSPwch1.0 than in the SHRSP/Izm. In addition, the ACT response to increases in SAP was stronger in the SHRSPwch1.0 than in the SHRSP/Izm. However, the ACT response to increases in HR did not differ between the SHRSPwch1.0 and SHRSP/Izm. Conclusions: These findings suggest that ACT continues for longer in SHRSPwch1.0 than in SHRSP/Izm. In SHRSPwch1.0, ACT is strongly correlated with the magnitude of increases in SAP, but not the SAP level itself or the size of increases in HR.

Exaggerated Stress Responses of a Chromosome-1 Congenic Strain Derived from Shrsp/Izm and Wky/Izm

P136 Background: QTL mapping studies showed that the rat chromosome 1 includes a gene responsible for hypertension in SHR. To understand roles of this putative hypertension gene on the cardiovascular physiology, we studied stress responses of a congenic strain with the chromosome-1 blood pressure (BP) QTL transferred from SHRSP/Izm. Methods: The region between D1Wox29 and D1Rat112 (∼60cM) was introgressed from SHRSP/Izm to WKY/Izm by repeated backcrossing. The established congenic strain (CONG) covered a 100:1 support interval for the BP QTL found in the previous study (Am J Hypert 1999;12:1098). After telemetry probes were implanted, age-matched male CONG and WKY (5-6 month old) were used in the following experiments; 1) Restraint stress: rats were put in small tubes for 1h at room temperature and BP was monitored with radiotelemetry. 2) Cold stress: rats were kept at 4 C for 2h and BP was monitored using telemetry. Results: Without the stresses, BP measured with telemetry was not different between WKY and CONG (Table) although tail-cuff measurement gave a significant difference in BP between the two strains (141±3.5 and 154±8.3mmHg for WKY and CONG, p