Elsevier

Neuropharmacology

Volume 50, Issue 2, February 2006, Pages 191-208
Neuropharmacology

Changes in purinergic signalling in developing and ageing rat tail artery: Importance for temperature control

https://doi.org/10.1016/j.neuropharm.2005.08.019Get rights and content

Abstract

This study aimed to examine the expression and function of P2 receptors of the rat tail and mesenteric arteries during maturation and ageing (4, 6 and 12 weeks, 8 and 24 months). Functional studies and receptor expression by immunohistochemistry revealed a heterogeneous phenotype of P2 receptor subtypes depending on artery age. The purinergic component of nerve-mediated responses in the tail artery was greater in younger animals; similarly responses to ATP and α,β-meATP and the expression of P2X1 receptors decreased with age. Contractile responses to 2-MeSADP decreased with age, and were absent at 8 and 24 months; P2Y1 receptor expression followed this pattern. UTP-induced contractions and P2Y2 receptor expression also decreased with age. The mesenteric artery contracted to UTP, responses at 4 and 6 weeks were larger than at other ages although P2Y2 receptor expression did not significantly differ with age. 2-MeSADP induced relaxation of the mesenteric artery, responses being greatest at 6 weeks and decreased thereafter, which was mimicked by the P2Y1 receptor immunostaining. We speculate that the dramatic changes in expression of P2 receptors in the rat tail artery, compared to the mesenteric artery, during development and ageing are related to the role of the tail artery in temperature regulation.

Introduction

It is now recognised that extracellular purines (adenosine, adenosine diphosphate (ADP) and adenosine 5′-triphosphate (ATP)) and pyrimidines (uridine diphosphate (UDP) and uridine 5′-triphosphate (UTP)) are important extracellular signalling molecules that mediate diverse effects on many biological processes (see Ralevic and Burnstock, 1998, Burnstock and Knight, 2004). Currently, seven mammalian P2X ionotropic ligand-gated ion channel receptors (P2X1–7) and eight P2Y metabotropic G protein-coupled receptors (P2Y1, P2Y2, P2Y4, P2Y6, P2Y11, P2Y12, P2Y13 and P2Y14) have been identified (Burnstock, 2004).

There is much evidence showing the existence of sympathetic purinergic co-transmission involving noradrenaline (NA) and ATP in a variety of blood vessels, including the tail artery of the rat (Sneddon and Burnstock, 1985) and the mesenteric artery of the guinea pig and rat (Ishikawa, 1985, Sjöblom-Widfeldt, 1990). The sympathetic nerves supplying different blood vessels release a variable ratio of NA:ATP, the purinergic component being relatively minor (∼10%) in the rat tail artery (Bao et al., 1993), whereas in the rabbit splenic artery, nerve-mediated responses are largely purinergic (Ren and Burnstock, 1997). In the rabbit mesenteric artery ATP is the sole excitatory transmitter, although NA is released and contributes to feedback inhibition (Ramme et al., 1987).

P2 receptors are widely distributed in the cardiovascular system and are important in the regulation of vascular tone (Boarder and Hourani, 1998). The P2X1 receptor is the principal P2X receptor subtype expressed on most vascular smooth muscles (Valera et al., 1994, Collo et al., 1996) and Bo and Burnstock (1993) reported that the smooth muscle of the rat tail artery has a high density of P2X1 receptors.

P2X2 receptor immunoreactivity has been visualised on smooth muscle cells of rat mesenteric, renal and pulmonary arteries, although at a lower density than P2X1 (Hansen et al., 1999). In addition, P2X4 receptors have also been visualised in rat aorta and vena cava as well as coronary, pulmonary, renal and femoral arteries (Soto et al., 1996, Nori et al., 1998).

Metabotropic P2Y receptors also participate in the control of vascular tone. At least four P2Y receptor subtypes mediate the vascular effects of extracellular nucleotides, namely P2Y1, P2Y2, P2Y4 and P2Y6 receptors. P2Y1 receptors are activated by the endogenous ligands ADP and ATP; in contrast P2Y2, P2Y4 and P2Y6 receptors are activated by uridine nucleotides. UTP was found to be equipotent with ATP at evoking contraction of the tail artery (Evans and Kennedy, 1994), suggestive of two populations of P2 receptors (McLaren et al., 1998), the second population being G protein-coupled P2Y2 and P2Y4 receptors.

P2Y1 receptors are distributed on both vascular smooth muscle and endothelium. When present on the endothelium they mediate vasodilatation via endothelial nitric oxide (NO; Moncada et al., 1991) and also by the generation of endothelium-dependent hyperpolarising factor (EDHF: Malmsjö et al., 1999, Stanford et al., 2001). Smooth muscle P2Y1 receptors also produce vasodilatation of a number of blood vessels, including the rabbit mesenteric artery (Mathieson and Burnstock, 1985, Burnstock and Warland, 1987) and vasoconstriction of others (Knight et al., 2003, Steinmetz et al., 2003). Similarly P2Y2 receptors in the vasculature are either present on the endothelium, such as in the rat and golden hamster isolated mesenteric arterial beds (Ralevic and Burnstock, 1996a, Ralevic and Burnstock, 1996b), where they mediate vasodilatation via NO synthesis and release, or on smooth muscle as in the rat and bovine middle cerebral artery (Miyagi et al., 1996) where they mediate contraction. In the rat tail and femoral arteries, UTP has been shown to produce vasoconstriction (Saïag et al., 1990).

Ageing produces changes in vascular smooth muscle cells and endothelial cells (Wei, 1992). These changes include thickening of the media and enlargement of the lumen diameter, increased stiffening and an accompanying reduction in arterial distensibility of larger vessels (see Moreau et al., 1998, Laurant et al., 2004). Generally, there is a reduced tendency for endothelium-dependent relaxation and endothelial release of NO with increasing age (Hynes and Duckles, 1987). Relaxations to histamine decline during ageing in the rat mesenteric artery (Moritoki et al., 1986), as do relaxations to ACh, partly due to reduced hyperpolarisation by EDHF (Marín, 1995). Endothelial-dependent relaxations to ATP and acetylcholine (ACh) are impaired in carotid arteries from old and hypertensive rats (Hongo et al., 1988). Endothelium-independent vasodilatations are also modified with age (Docherty, 1990, Marín, 1995).

There is a reduction during ageing in the maximum contractile responses induced by NA in rat, rabbit and guinea-pig aortae, dog and monkey mesenteric arteries, dog cerebral arteries and rat tail arteries (Fouda and Atkinson, 1986, Marín, 1995). In contrast, contractions to NA are increased with age in isolated dog iliac, carotid, renal and mesenteric arteries (Cox et al., 1976), ear artery from lambs and ewes (Wyse et al., 1977) and rat aorta (Olah and Rahwan, 1987). The isolated rabbit aorta (Hayashi and Toda, 1978) and rat femoral and carotid arteries (Duckles et al., 1985) are unaffected by increasing age.

Considerable plasticity during development and ageing of the autonomic nervous system in terms of co-transmitter expression and function is apparent. Hormones, disease, surgery and trauma can also underlie plasticity in the autonomic nervous system (Burnstock, 1997). Changes in the structure of neurons and nerves (Cowen, 1993) and on adrenergic neurotransmission in the sympathetic nervous system occur with age. Neuron structure can change, as well as a reduction in neuron number and loss of nerve fibres. Dhall et al. (1986) showed developmental changes in perivascular noradrenergic and peptide-containing nerves in the guinea pig; noradrenergic nerve density peaked 4 weeks after birth whilst nerves containing VIP, CGRP and SP peaked at birth and declined thereafter in the mesenteric and carotid arteries.

The rat tail artery is important for both balance and thermoregulation. Body appendages, such as the tail, are important heat exchange sites where increases in blood flow cause heat dissipation. Skin blood flow results in heat loss from the surface and this is modified by a sympathetically mediated mechanism. The tail of the rat functions as a heat-loss organ since it lacks hair covering, has a large surface area to volume ratio and is highly vascularised with arteriovenous anastomoses (Gordon, 1990). Up to 20% of total body heat loss can occur via the tail.

The aim of this study was to examine the expression and function of P2 receptors in the rat tail and mesenteric arteries during maturation and ageing. Rats attain sexual maturity at 2–3 months of age; they show rapid growth up to 8–9 months, which becomes stable by about 12 months. Twenty-four months is a commonly accepted age for senescence in rats (Weihe, 1987). For this study young rats of 4 and 6 weeks old were used to show any changes occurring in the early life of a rat, 12-week-old rats were used to reflect any changes as sexual maturity was reached, 8-month-old adult rats whose rapid growth was beginning to slow down and old (senescent) rats aged 24 months were used.

Section snippets

General procedures

Male Sprague–Dawley rats, aged 4, 6 and 12 weeks, 8 and 24 months, were killed according to Home Office (UK) regulations covering Schedule one procedures–by asphyxiation with CO2 and subsequent cervical dislocation to confirm death. The tail and mesenteric arteries were dissected free, carefully cleaned of adhering fat and excessive connective tissue. The arteries were cut into rings approximately 4 mm in length and mounted horizontally for tension recording by inserting two tungsten wires through

Responses to nerve stimulation in the tail artery

EFS of the tail arteries from each age group induced sympathetic nerve-mediated, frequency-dependent contractions that were TTX (1 μM) sensitive. In the presence of suramin (100 μM) alone (Fig. 1a–e), contractile responses induced by EFS in all age groups were significantly inhibited (P < 0.05 or less). When the frequency–response curves were repeated in the presence of suramin and prazosin (1 μM) contractions were almost completely inhibited for each age group (P < 0.001). Conversely, when the curves

Discussion

This study has shown that the function and expression of P2X and P2Y receptors of the rat tail and mesenteric artery alter with maturation and age. In general the trend is for the sensitivity and expression of P2X and P2Y receptors to decrease as the age of the animal increases. However, there were significant differences between the two vessels examined that include transient maturational increases in the expression of selected receptors in the mesenteric artery.

Bao et al. (1993) previously

Acknowledgements

The authors thank Dr H.-Z. Ruan for invaluable assistance with immunohistochemistry, and Mr Tim Robson for excellent technical assistance.

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