Endogenous angiotensin II-induced p44/42 mitogen-activated protein kinase activation mediates sodium appetite but not thirst or neurohypophysial secretion in male rats

Abstract

The renin-angiotensin-aldosterone system makes a critical contribution to body fluid homeostasis, and abnormalities in this endocrine system have been implicated in certain forms of hypertension. The peptide hormone angiotensin II (AngII) regulates hydromineral homeostasis and blood pressure by acting on both peripheral and brain targets. In the brain, AngII binds to the angiotensin type 1 receptor (AT1R) to stimulate thirst, sodium appetite and both arginine vasopressin (AVP) and oxytocin (OT) secretion. The present study used an experimental model of endogenous AngII to examine the role of p44/42 mitogen-activated protein kinase (MAPK) as a signalling mechanism to mediate these responses. Animals were given a combined treatment of furosemide and a low dose of captopril (furo/cap), comprising a diuretic and an angiotensin-converting enzyme inhibitor, respectively, to elevate endogenous AngII levels in the brain. Furo/cap induced p44/42 MAPK activation in key brain areas that express AT1R, and this effect was reduced with either a centrally administered AT1R antagonist (irbesartan) or a p44/42 MAPK inhibitor (U0126). Additionally, furo/cap treatment elicited water and sodium intake, and irbesartan markedly reduced both of these behaviours. Central injection of U0126 markedly attenuated furo/cap-induced sodium intake but not water intake. Furthermore, p44/42 MAPK signalling was not necessary for either furo/cap- or exogenous AngII-induced AVP or OT release. Taken together, these results indicate that p44/42 MAPK is required for AngII-induced sodium appetite but not thirst or neurohypophysial secretion. This result may allow for the discovery of more specific downstream targets of p44/42 MAPK to curb sodium appetite, known to exacerbate hypertension, at the same time as leaving thirst and neurohypophysial hormone secretion undisturbed.

Figure 1

Drawing of a coronal hemi section of rat brain depicting the brain regions of interest for p44/42 MAPK activation assays. Circles within each brain region represent 1 mm micropunches collected for the p44/42 MAPK ELISAs. The brain regions examined include circumventricular organs OVLT and SFO, as well as the PVN and SON of the hypothalamus. Additionally, the S1 was collected as a control brain region to assay P44/42 MAPK activation. Abbreviations: S1 = primary somatosensory cortex, CPu = caudate putamen, MS = medial septal nucleus, OVLT = organum vasculosum of the lateral terminalis, SFO = subfornical organ, fi = fimbria of the hippocampus, ic = internal capsule, PVN = paraventricular nucleus of the hypothalamus, SON = supraoptic nucleus of the hypothalamus.

Figure 2

Bar graphs illustrating the levels of total p44/42 MAPK and phospho p44/42 MAPK in the OVLT/SFO (panel A), PVN/SON (panel B), and S1 (panel C) after either vehicle or furo/cap treatment sc and either vehicle, irbesartan, or U0126 administered icv (n = 3-5/group). Total p44/42 MAPK levels do not change, whereas furo/cap treatment increases phosphorylated p44/42 MAPK in the OVLT/SFO and PVN/SON, but not in S1. Both irbesartan and U0126 prevent this furo/cap induced increase in p44/42 MAPK phosphorylation. Abbreviations: OVLT = organum vasculosum of the lateral terminalis, SFO = subfornical organ, PVN = paraventricular nucleus of the hypothalamus, SON = supraoptic nucleus of the hypothalamus, S1 = primary somatosensory cortex.

Figure 3

Line graphs illustrating the effect of furo/cap treatment on urine volume (panel A) and water and sodium intake during a two-bottle test (panel B). On the x axis, zero minutes represents the time of captopril injection. Compared with vehicle treatment, rats given furo/cap treatment excreted significantly more urine within 15 minutes (n=16). Furo/Cap also induced a robust intake of water and 1.5% sodium within 45 minutes and 60 minutes of treatment, respectively. Asterisks indicate that p<0.01 when comparing Furo/Cap to vehicle treatment.

Figure 4

Line graphs illustrating the effect of blocking AT1R receptors and MAPK activation after furo/cap treatment in a two bottle test (panel A) and in a one bottle test (panel B). On the x axis, zero minutes represents the time of captopril injection. In the two bottle test, compared with Furo/Cap + Vehicle treatment, rats given the Furo/Cap + AT1R antagonist irbesartan treatment consumed significantly less water within 45 minutes after treatment (n=28). Irbesartan also reduced the intake of 1.5% saline within 45 minutes of treatment. Compared with Furo/Cap + vehicle treatment, rats given Furo/Cap + the MEK inhibitor U0126 treatment consumed significantly less water within 90 minutes after treatment and significantly less saline within 30 minutes of treatment (n=28). In the one bottle test, compared with vehicle treated, rats given the MEK inhibitor U0126 treatment consumed similar amounts of water (n=12). Asterisks indicate that p<0.05 when comparing to Furo/Cap + Veh treatment. Stats for Furo/Cap + Veh compared to Veh are shown in Figure 3.

Figure 5

Bar graphs illustrating the role of p44/42 MAPK activation on Furo/Cap-induced neurohypophysial secretion. Plasma AVP (Panel A) and OT (Panel B) after vehicle or Furo/Cap treatment combined with either vehicle, irbesartan, or U0126 administered icv (n= 3-5/group). Furo/Cap-induced AVP and OT secretion was not diminished with either the AT1R antagonist irbesartan or the MEK inhibitor U0126. Abbreviations: AVP = arginine vasopressin, OT = oxytocin.

Fig 6

Bar graphs illustrating the effect of inhibiting p44/42 MAPK activation on AngII-induced neurohypophysial hormone secretion. Plasma AVP (Panel A) and OT (Panel B) levels two and fifteen minutes after icv Veh, AngII, or U0126 plus AngII (n = 10-16/group). AngII-induced AVP and OT levels were not inhibited with the MEK inhibitor U0126. Abbreviations: AVP = arginine vasopressin, OT = oxytocin.