Proceedings of The Physiological Society
University College Dublin (2009) Proc Physiol Soc 15, C41
Are the projections from the periaqueductal grey to pontine noradrenergic neurones excitatory or inhibitory?
L. Hickey1, E. Sher2, B. Lumb1, A. E. Pickering1
1. Department of Physiology & Pharmacology, University of Bristol, Bristol, United Kingdom. 2. Lilly Research Centre, Eli Lilly, Windlesham, Surrey, United Kingdom.
The periaqueductal grey (PAG) is a midbrain site involved in the modulation of nociception and is believed to control nociception, in part, by activating pontospinal noradrenergic (NA) neurones. Connections between the PAG and noradrenergic cell groups have previously been demonstrated (Bajic et al., 2001), however, the neurotransmitter phenotype is as yet unknown. To address this issue we have injected viral vectors to the ventrolateral PAG (VL-PAG) to anterogradely label projections to pontine noradrenergic (NA) neurones in combination with immunocytochemistry for glutamate (vGlut1&2) and GABA (vGat) transporters. Male Wistar rats (n=3) received injections of AAV-CMV-EGFP (250nl) under anaesthesia (ketamine 60mg.kg-1/medetomidine 25µg.kg-1 i.p) into the VL column of the PAG. After 8 days all animals were terminally anaesthetised with sodium pentobarbital (70mg.kg-1 i.p.) and perfusion fixed with 4% formalin, brains were removed and 40μm sections were cut. Sections were processed immunocytochemically to identify dopamine β-hydroxylase and visualise terminals containing EGFP and VGlut1&2 or VGat. Injection sites and terminal labelling were determined using confocal imaging and 3D reconstruction software (Volocity 4, Improvision). AAV-CMV-EGFP produced strong terminal labelling in the pons with a predominantly ipsilateral distribution. Terminals showing co-localisation with Vglut or Vgat were identified in A5, A6 and A7 cell groups indicating excitatory or inhibitory amino acid (EAA or IAA) projections from the VL-PAG. Quantification of terminals closely apposing NA cell bodies in these regions showed that the LC received the densest innervation of EAA and IAA profiles. However after analysis of the density of innervation per NA neuron, the A7 cell group received the largest number of EAA and IAA profiles per soma (3.5±1.8 and 3.8±1.5 profiles/soma respectively). By contrast the A6 cell group received moderate EAA and IAA innervation (1.3±0.4 and 1.1±0.3 profiles/soma). The A5 territory received more IAA (3.5±2.2 profiles/soma) than EAA innervation from the VL PAG (1.5±1.0 profiles/soma). In conclusion, we show that VL PAG projections to A5, A6 and A7 have terminals closely apposed to NA neurones. These terminals express vesicular transporters for EAA and IAA suggesting that there are both excitatory and inhibitory phenotypes. The A7 region is densely innervated by glutamatergic and GABAergic profiles, which may identify the neurotransmitter phenotype of previously documented symmetrical and asymmetrical synapses made by PAG projections to A7 (Bajic et al 2001). We have also shown that the A5 region receives a strong inhibitory input from the PAG, which may be responsible for the depressor response that occurs on activation of VL PAG. These data provide evidence for a specific and differential control of pontine NA neurones by the VL PAG.
Where applicable, experiments conform with Society ethical requirements