Lipopolysaccharide (LPS)-induced hypotension in ramp2 overexpressing mice

* NC Clark, K Husmann, JA Fischer, W Born, *M Connolly & *SD Brain, *Cardiovascular Division, King’s College London, SE1 1UL; Research Laboratory for Calcium Metabolism, Department of Orthopaedic Surgery Balgrist, University of Zurich, Switzerland.

Adrenomedullin1-52 (AM) is a peptide of the calcitonin family. AM receptor subtypes, AM1 and AM2, are composed of the G protein-linked calcitonin receptor-like receptor (CLR) and receptor activity modifying protein (RAMP) 2 or -3, respectively (McLatchie, 1998). Circulating AM is elevated in sepsis, but its role and the receptors involved, especially in sepsis-induced hypotension, are unclear. We sought to investigate the effects of overexpression of RAMP2 in an experimental LPS model of sepsis in mice.

Heterozygous transgenic (TG) C57Bl/6 mice that overexpress RAMP2 under control of a smooth muscle α-actin promoter in corresponding tissues, e.g. the blood vessels, were compared with wild-type (WT) littermates. The mice exhibited normal growth and similar baseline measurements for mean arterial pressure (MAP: WT=112 ± 7, TG=114 ± 4, mm Hg, mean ± s.e.mean, n=15-16) and heart rate (HR: WT=703 ± 16, TG=715 ± 18, n=15-16), as assessed with the tail cuff method. This suggests that RAMP2 is not a limiting factor for an effect of AM on basal blood pressure or heart rate.

The hypotensive effect of AM (130pmol/25g in 100 µl/25g, administered via tail vein) was assessed in conscious, female, age-matched WT and RAMP2 TG mice with the tail cuff method. TG mice displayed a significantly (p<0.01) greater hypotensive response to AM (% change from pre-injection: WT=6 ± 6, n=6; TG=36 ± 7, n=6). These studies demonstrate that the RAMP2 TG mice possess increased levels of functional CLR/RAMP2 receptor complexes and that AM1 receptors can mediate AM-induced hypotension.

In WT and TG mice matched for age and gender, blood pressure (BP) was assessed before and 1.5 and 4 h after subcutaneous abdominal injection of LPS (5mg and 10ml/kg, serotype: 0127:B8) or vehicle (saline, 10ml/kg). After 1.5 h, LPS produced a similar (p=0.32) fall in BP in WT (% change 25±10, n=8) and TG (% change 37±5, n=7) mice. However, the fall in BP compared to vehicle was significant (p=0.01) in TG mice, but not in WT mice (p=0.06). At 4 h after LPS treatment, BP had fallen further in WT mice (% change 39±3, n=7), but stabilised in the TG (% change 36±6, n=6) mice. Again the responses to LPS in WT and TG mice were indistiguishable (p=0.77). At this time point responses were significantly different compared to the respective vehicle responses in both WT (p<0.0001) and TG (p=0.0144) mice.

Taken together, the data demonstrate that exogenous AM induces enhanced hypotensive responses in RAMP2 TG as compared to WT mice. Furthermore, overexpression of RAMP2 and hence of AM1 receptors may slightly accelerate the development of sepsis-induced hypotension, but has no effect on the ultimate severity of the hypotensive response observed in this study.

McLatchie,L.M et al., (1998) Nature 393 p333.

NC Clark is a BHF Ph.D. student.