Elsevier

Autonomic Neuroscience

Volume 86, Issues 1–2, 28 December 2000, Pages 18-29
Autonomic Neuroscience

Cotransmission from sympathetic vasoconstrictor neurons: differences in guinea-pig mesenteric artery and vein

https://doi.org/10.1016/S1566-0702(00)00203-4Get rights and content

Abstract

Vasoconstrictor responses to electrical field stimulation (EFS, 0.2–32 Hz, 0.1 ms, 12 V, for 1 min) were measured in endothelium-denuded segments of guinea-pig mesenteric vein and compared to responses in mesenteric artery. The distribution of both tyrosine-hydroxylase-like immunoreactivity (TH-LI) and neuropeptide Y-like immunoreactivity (NPY-LI) was also studied using anti-TH and anti-NPY antibodies. The effect of exogenous NPY (10 nM) on EFS (8 Hz, 0.3 ms, 12 V, for 1 min)-evoked overflow of noradrenaline (NA) was also studied using an HPLC technique with electrochemical detection. Veins responded with contractions at lower frequencies of stimulation than arteries. Prazosin (0.1 μM) abolished the EFS-evoked contractions in artery at 0.5–32 Hz and in vein at 0.2–1 Hz of stimulation. However, in vein, the contractile responses to EFS at 2–32 Hz of stimulation were only reduced by prazosin. Phentolamine (1 μM) abolished the responses to 0.5–4 Hz and reduced the responses to 8–32 Hz of EFS in artery. In vein, phentolamine (1 μM) abolished the responses to 0.2–1 Hz and facilitated the contractions elicited by 16–32 Hz. The NPY-receptor antagonist BIBP3226 (1 μM), in combination with phentolamine, abolished contractions in vein. Yohimbine (0.1 μM) abolished the responses to lower frequencies of stimulation in both artery (0.5–2 Hz) and vein (0.2–1 Hz). The responses to greater frequency stimulation were not affected by yohimbine in artery, and were facilitated in vein. Pre-treatment of animals for 24 h with reserpine abolished contractile responses to EFS in artery, whereas in vein, responses to 0.2–2 Hz were abolished while responses to 4–32 Hz were unchanged. Suramin (100 μM) or α,β-methylene ATP (α,βMeATP; 10–100 μM) treatment did not affect the contractile responses to EFS in either artery or vein. Pyridoxal-phosphate-6-azophenyl-2′,4′-disulfonic acid tetrasodium (PPADS; 30 μM), even potentiated the responses to 2–16 Hz in vein. However, following reserpine-treatment, both PPADS and suramin reduced the nerve-evoked contractions of vein. Either BIBP3226 (1 μM) alone or BIBP3226 in combination with PPADS or suramin abolished the contractile response to EFS in reserpine-treated veins. NPY (100 nM) produced significantly more contraction in vein than in artery (i.e., 93±2.5 versus 7±4% of the response to 70 mM KCl, respectively). NPY (10 nM) significantly reduced the NA overflow evoked by EFS at 8 Hz. Flat mount preparations and cryostat sections of both mesenteric artery and vein revealed that TH-LI and NPY-LI were co-localized in a dense network of fibers within the adventitial layer. In conclusion, NA exclusively mediates the contractile response to sympathetic nerve stimulation in guinea-pig mesenteric artery, whereas at least three neurotransmitters [i.e., NA, adenosine 5′-triphosphate (ATP) and NPY] are involved in the neural response of mesenteric vein.

Introduction

There is growing appreciation that peripheral nerves release more than one transmitter, a process referred to as plurichemical neurotransmission or cotransmission (Furness et al., 1989, Burnstock, 1990). The evidence for cotransmission is particularly strong for postganglionic sympathetic nerves, which have been shown to corelease adenosine 5′-triphosphate (ATP), neuropeptide Y (NPY) and noradrenaline (NA) (for reviews, see Stjärne, 1989, Burnstock, 1990, Westfall et al., 1991, Lundberg, 1996). However, the relative contribution of each of these neurotransmitters to contraction varies significantly between blood vessels. For example, both NA and ATP significantly contribute to sympathetic neural responses in rabbit saphenous and jejunal artery, and in guinea-pig submucosal arterioles (see Burnstock, 1995). In contrast, it is more difficult to reveal a nonadrenergic component of nerve stimulation in rat tail artery (see Burnstock, 1995), even though ATP is released during nerve stimulation in this vessel (Msghina et al., 1992, Mutafova-Yambolieva and Westfall, 1998).

Both arteries and veins contain smooth muscle and both are innervated by sympathetic nerves. However, the physiological roles of high-pressure arteries versus low-pressure veins differ significantly from one another, i.e., in the splanchnic circulation, arteries and arterioles contribute to total peripheral resistance and control the local flow of blood to the gastrointestinal tract. In contrast, the venous side of the splanchnic circulation has a storage capacity and, hence, contributes to the regulation of blood volume and venous return, a role referred to as capacitance. Thus, sympathetic nerves have been shown to play significantly different roles in controlling the arterial and venous sides of the splanchnic circulation (see Hansen et al., 1998).

The goal of the present study was to investigate cotransmission in the splanchnic circulation by comparing vessels taken from the two functionally distinct portions of this vascular bed, i.e., mesenteric arteries and veins.

The electrical and contractile response elicited with stimulation of sympathetic nerves has been shown to differ significantly between arteries and veins. In arteries (including the guinea-pig mesenteric artery), sympathetic nerve stimulation gives rise to rapidly rising excitatory junction potentials (EJPs) of brief duration that are resistant to prazosin, but are blocked by α,β-methylene ATP (α,βMeATP) (see Brock and Cunnane, 1992, Kreulen, 1986, Hottenstein and Kreulen, 1987). ATP is considered to be a prime candidate to mediate these EJPs (Burnstock, 1990, Stjärne et al., 1994). In contrast to arteries, fast EJPs are generally not observed with sympathetic nerve stimulation in veins (including the guinea-pig mesenteric vein). Rather, repetitive nerve stimulation in vein typically produces a slow depolarization (Suzuki, 1981) that is resistant to purinoceptor desensitization with α,βMeATP (Hottenstein and Kreulen, 1987, Hirst and Jobling, 1989). This has led to the conclusion that ATP is not involved in these junctional events (Hottenstein and Kreulen, 1987, Hirst and Jobling, 1989) and, hence, is not released during nerve stimulation. However, in a study measuring ATP release in vein (i.e., human saphenous vein), the conflicting observation was made that ATP was released together with NA but did not appear to generate the non-adrenergic component of contraction (Rump and von Kügelgen, 1994). Therefore, differences in the mechanical and electrical responses to nerve stimulation between arteries and veins might be accounted for by differences in the end (i.e. postjunctional) effects of released neurotransmitters.

NPY is considered to be a cotransmitter, together with NA and ATP, in the sympathetic nervous system (Lundberg et al., 1990). This peptide is assumed to predominantly mediate responses to higher frequencies of stimulation and therefore play a role specifically at high levels of sympathetic nerve activity (see Lundberg, 1996). NPY is known to cause contraction of the vascular smooth muscles in some regions, such as rat mesenteric arterial bed, human coronary artery, cerebral circulation (Abounader et al., 1995, Macho et al., 1987, Westfall et al., 1987). In some systems, NPY can also play a role as neuromodulator, by potentiating the vasoconstriction produced by other agonists, or by sympathetic nerve stimulation (Cortes et al., 1999) or inhibiting the release of NA (Westfall et al., 1987). The role of this peptide in the mesenteric artery and vein of guinea-pig is unclear.

In the present study, the relative contribution of NA, ATP and NPY to neurally induced contractions in guinea-pig mesenteric artery and vein was established. Autonomic neurotransmission was further characterized morphologically by comparing immunohistochemically the localization of tyrosine-hydroxylase-like immunoreactivity (TH-LI) and NPY-like immunoreactivity (NPY-LI) in artery and vein.

Section snippets

General

Male guinea-pigs (weighing 400 to 450 g) were sacrificed by CO2 overdose followed by exsanguination, in keeping with protocols approved by the University of Nevada’s Animal Care and Use Committee. The aorta, inferior mesenteric artery, vein and associated mesentery were immediately removed from the animal and placed in cold (10°C) oxygenated Krebs solution for further dissection of the mesenteric blood vessels. Segments of first- and second-order branches of the inferior mesenteric artery

Results

The contractile responses to electrical field stimulation (EFS, 0.2–32 Hz) were measured in artery and vein and compared to the response obtained with 70 mM KCl. KCl produced a significantly greater contraction in artery than vein (i.e., 110.25±13.5 mN/mm2 in artery versus 71.4±5.9 mN/min.mm2 in vein, P<0.05). These responses to 70 mM KCl are submaximal in both the artery and vein, i.e., they represent approximately 70% of the maximum contraction developed in response to combined application of

Discussion

Guinea-pig mesenteric artery and vein exhibit significant differences with regard to their electrical and mechanical responses to nerve stimulation (Kreulen, 1986, Hottenstein and Kreulen, 1987, Hirst and Jobling, 1989, Browning et al., 1999). The present study generally confirms these earlier observations and also reveals substantial differences in the role of sympathetic co-transmission in these two vascular networks. Our results suggest that NA is the primary excitatory neurotransmitter in

Acknowledgements

This work was supported by US Public Health Service Grant HL 60031 to V.M-Y.

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