HYPERPOLARIZATION BY OPIOIDS ACTING ON MU-RECEPTORS OF A SUBPOPULATION OF RAT PERIAQUEDUCTAL GRAY NEURONS IN-VITRO

1 The actions of opioids on membrane properties of rat periaqueductal gray neurones were investigated using intracellular recordings from si ngle neurones in brain slices. Morphological properties and anatomical location of each impaled neurone were characterized by use of intrace llular staining with biocytin. The present paper primarily considers n eurones which were directly hyperpolarized by opioids. The accompanyin g paper considers inhibition of synaptic transmission by opioids. 2 Me t-enkephalin (10-30 mu M) hyperpolarized 29% (38/130) of neurones. The hyperpolarization was fully antagonised by naloxone (1 mu M, n = 3). The response to Met-enkephalin was not affected by agents which block synaptic neurotransmission (1 mu M tetrodotoxin, and 0.1 mu M tetrodot oxin + 4 mM Co2+, n = 3). 3 The specific mu-receptor agonist, D-ala-me t-enkephalin-glyol (3 mu M, n = 17) produced hyperpolarizations of sim ilar amplitude to those produced by Met-enkephalin (10-30 mu M). The E C(50) of D-ala-metenkephalin-glyol was 80 nM and the maximum response was achieved at 1-3 mu M. The delta-receptor (D-Pen-D-Pen-enkephalin, 3 mu M, n = 7) and kappa-receptor (U50488H, 3 mu M, n = 5) agonists ha d no effect on the membrane properties of these neurones. 4 The opioid -induced hyperpolarization was associated with an increased potassium conductance. Hyperpolarizations were accompanied by a significant decr ease in membrane resistance between -70 and - 80 mV, and a significant ly greater decrease between - 110 and - 140 mV (n = 16). Hyperpolariza tions reversed polarity at -111 +/- 3 mV (n = 16), close to the expect ed equilibrium potential for potassium ions. The reversal potential of outward currents increased by 24 mV when the extracellular potassium concentration was raised from 2.5 to 6.5 mM, which is close to the val ue predicted by the Nernst equation (25 mV) for a potassium conductanc e. 5 Resting inward rectification (reduced input resistance at potenti als more negative than - 100 mV in the absence of opioids) was signifi cantly greater in neurones which were hyperpolarized by opioids than i n those which were not hyperpolarized. The amplitude of action potenti al after hyperpolarizations was significantly smaller in neurones whic h were hyperpolarized by opioids. Other membrane properties did not di ffer significantly between opioid-sensitive and -insensitive neurones. 6 Neurones hyperpolarized by opioids were multipolar (58%), triangula r (21%) or fusiform (5%) in shape with a soma diameter of 22 +/- 1 mu m (n = 19, longest axis). Dendritic spread was in a large radiating pa ttern, usually in all directions, with axons usually originating from primary dendrites. The axons were usually branched and projected in se veral directions. Morphological properties did not differ significantl y between opioid-sensitive and -insensitive neurones. 7 Neurones hyper polarized by opioids were located predominantly in the lateral periaqu eductal gray, as well as in the more dorsal areas of the ventrolateral periaqueductal gray, whereas neurones not hyperpolarized by opioids w ere located in the more ventral areas of the ventrolateral periaqueduc tal gray. 8 These studies demonstrate that opioids acting on mu-recept ors increase potassium conductance in a sub-population of large neuron es located predominantly in the lateral column of the periaqueductal g ray. The neurones hyperpolarized by opioids could be involved in the a ntinociceptive actions of opioids, but might also be involved in other functions because a large proportion lie outside of the main 'antinoc iceptive zone' of the periaqueductal gray. It is also unlikely that th ese neurones are GABAergic, suggesting that they might not participate in the postulated antinociceptive action of opioids mediated via disi nhibition of neurones which project to the ventral medulla.