Enhanced hemeoxygenase activity in the rostral ventrolateral medulla mediates exaggerated hemin-evoked hypotension in the spontaneously hypertensive rat

Abstract

In anesthetized normotensive rats, activation of brainstem hemeoxygenase (HO) elicits sympathoinhibition and hypotension. Accordingly, we tested the hypothesis that attenuated basal or induced HO activity in the rostral ventrolateral medulla (RVLM) contributes to hypertension in the spontaneously hypertensive rat (SHR). We measured basal RVLM HO expression and catalytic activity and investigated the effects of intra-RVLM HO activation (hemin) or selective HO isoform 1 (HO-1) inhibition [zinc protoporphyrin IX (ZnPPIX)] on mean arterial pressure (MAP), heart rate, and RVLM neuronal norepinephrine (NE) level (index of sympathetic activity) in conscious SHRs and Wistar Kyoto rats. Basal RVLM HO catalytic activity (bilirubin level) and HO-1 expression were significantly higher in the SHR. These neurochemical findings were corroborated by the significantly greater decreases (hemin) and increases (ZnPPIX) in RVLM NE and MAP in the SHR. By contrast, HO-independent CO release in the RVLM (CO-releasing molecule 3) elicited similar MAP reductions in both rat strains. Furthermore, pretreatment with ZnPPIX or the selective neuronal nitric-oxide synthase (nNOS) inhibitor N-propyl-l-arginine abrogated the neurochemical (RVLM cGMP) and hypotensive responses caused by hemin. In addition to demonstrating, for the first time, higher basal RVLM HO catalytic activity and HO-1 expression in the SHR, the findings suggest: 1) the exaggerated hypotension elicited by intra-RVLM HO activation in the SHR is nNOS-dependent, and 2) in the SHR, the enhanced RVLM HO-nNOS signaling compensates for the reduced expression/activity of the downstream target, soluble guanylyl cyclase. Together, the findings suggest a protective role for the RVLM HO-nNOS pathway against further increases in MAP in the SHR.

Fig. 1.

Time-course changes in RVLM NE (top) and MAP (bottom) in response to intra-RVLM hemin (1 nmol) microinjection in conscious SHRs and WKY rats. Values are means ± S.E.M. (n = 5 per group). *, P < 0.05 compared with corresponding values from WKY rats.

Fig. 2.

Time-course changes in RVLM NE (top) and MAP (bottom) after intra-RVLM microinjection of the selective HO-1 inhibitor ZnPPIX (1 nmol) and subsequent intra-RVLM hemin (1 nmol) microinjection in conscious SHRs and WKY rats. Values are means ± S.E.M. (n = 5 per group). *, P < 0.05 compared with corresponding values from WKY rats.

Fig. 3.

Bar graph depicting HO activity in control/treatment groups of SHRs and WKY rats. Values are means ± S.E.M (n = 5 per group). Basal RVLM HO activity is significantly (*, P < 0.05) higher in SHRs, and intra-RVLM hemin increased HO activity 4-fold in SHRs versus 2-fold in WKY rats compared with their respective aCSF levels (#, P < 0.05). The selective HO-1 inhibitor ZnPPIX abrogated (@, P < 0.05) the increases in HO activity caused by intra-RVLM hemin in both strains of rats.

Fig. 4.

Top, bar graph depicting the basal expression of the two sGC subunits, α1 and β1 subunits. Bottom, basal RVLM cGMP and its response to intra-RVLM hemin (1 nmol) in the absence or presence of ZnPPIX (1 nmol) or NPLA (12.5 μg/kg i.v.) in SHRs and WKY rats is shown. Values are means ± S.E.M (n = 4–5 per group). *, P < 0.05, compared with respective basal level; #, P < 0.05, compared with WKY rats.

Fig. 5.

Bar graphs showing expression of HO-1 (top), HO-2 (middle), and p-nNOS (bottom) in the RVLM of SHRs and WKY rats (n = 5 per group) in the absence (basal) or after intra-RVLM hemin (1 nmol). HO-1 and HO-2 were expressed in relation to actin, whereas p-nNOS was expressed in relation to total nNOS. *, P < 0.05, compared with corresponding value from WKY rats; #, P < 0.05, compared with untreated (basal) level.

Fig. 6.

Bar graph depicting RVLM NOx levels in control/treatment groups of SHRs and WKY rats. Values are means ± S.E.M (n = 5 per group). Hemin significantly (*, P < 0.05) increased RVLM NOx in SHR, and this response was abrogated by pretreatment with the selective nNOS inhibitor NPLA.

Fig. 7.

Time-course changes in MAP in SHRs (bottom) and WKY rats (top) pretreated with NPLA (12.5 μg/kg i.v) or an equal volume of saline. Intra-RVLM hemin (1 nmol) or equal volume of aCSF was microinjected, 30 min after NPLA/saline, into the RVLM of conscious SHRs or WKY rats. Values are means ± S.E.M. (n = 5 per group). *, P < 0.05, compared with corresponding control values.

Fig. 8.

Time-course changes in MAP elicited by intra-RVLM microinjection of CORM-3 in conscious SHRs and WKY rats. Values are means ± S.E.M. (n = 5 per group).