C.D. Barnes and 0. Pornpeiano (Eds.) Progress in Brain Research, Vol. 88 0 1991 Elsevier Science Publishers B.V.
24 1 CHAPTER 18
Responses of locus coeruleus neurons to neuropeptides H.-R. Olpe and M. Steinmann Research and Deaelopment Department, Pharmaceuticals Division, CIBA-GEIGY Ltd., Basel, Switzerland
The knowledge on the neuronal inputs to the locus coeruleus (LC) and their roles in regulating noradrenergic (NA) cellular activity is quite advanced. In recent years, however, about ten neuropeptides were found to be localized in the area of the rodent LC; peptides which may be considered as potential transmitters o r modulators acting in this area. Electrophysiological studies performed in viuo and in Liitro have revealed that many of these peptides are able to alter LC neuronal activity. Stimulatory effects have been described with vasopressin, substance P, adrenocorticotropin hormone and corticotropin-releasing factor. Depressant effects were seen with galanin, somatostatin, neuropeptide Y and enkephalin. Variable actions were observed in the case of neurotensin. While these findings point to a possible regulatory function of these peptides in this area, precise roles
remain unclear. Important information is lacking that would conclusively demonstrate their regulatory functions. It should be determined whether the stimulation of peptidergic cells elicits synaptic effects identical to the ones observed with local exogenous peptide applications. By studying the action of blockers of these transmitter and modulator candidates, we would probably begin to understand their importance in the regulation of tonic and phasic activity components. The LC is generally considered to consist of a homogenous group of neurons. T h e recent observation that subpopulations of these cells contain peptides as in the case of neuropeptide Y, galanin and vasopressin, points to the possible existence of subgroups of neurons having different functions.
Key words: locus coeruleus, neuropeptides, electrophysiology, modulator, rat
Introduction
The locus coeruleus (LC) of the rat brain is generally considered to be a homogenous and compact nucleus containing mainly noradrenergic (NA) neurons. Intra- and extracellular electrophysiological recordings are in keeping with this notion since the biophysical properties of all cells appear to be very similar (Williams et al., 1984). Over the past ten years, a steadily increasing number of potential physiological roles have been proposed that are claimed to be linked with this
brain area (Olpe et al., 1985; Svensson, 1987). We are, therefore, facing the problem that a small number of seemingly homogeneous neurons is thought to be involved in several distinct brain functions. One theoretical approach for solving this problem is to search for a common basic action of noradrenaline that would be coupled with a number of different, more complex functions. Alternatively, a basis for the heterogeneity of functions could be found in the fact that the LC projections have recently been shown to be selective and not as widely and diffusely spread
242
TABLE 1 Neuropeptides present in the area of the locus coeruleus Peptides present in locus coeruleus
Localized in perikarya
vasopressin substance P adrenocorticotropin hormone, ACTH corticotropin-releasing factor, CRF somatostatin. SRIF neuropeptide Y enkephalin neurotensin galanin delta sleep-inducing peptide, DSIP
as originally described (Grzanna et al., 1987). Another possible suggestion for the heterogeneity of LC neurons themselves derives from recent reports that several neuropeptides are present in this brain area which have been localized in fibers and/or cell bodies. A survey of the kind of peptides described in this area is provided in Table 1 (see also Sutin and Jacobowitz, this volume). It is remarkable that the rodent LC contains not only a considerable number of different peptides, but some of them are present at rather high concentrations, as in the case of substance P (Douglas et al., 1982) and somatostatin (Palkovits et al., 1982). The presence of these peptides raises the question of their physiological functions. Electrophysiological techniques provide a means of addressing this issue, at least partly, in that the sensitivity of LC neurons to these peptides can be investigated. While such studies give valuable information on the effects of peptides on cell activity, their membrane potentials and other biophysical membrane properties, they do not necessarily solve the problem of identifying the peptides’ physiological roles. Such information may eventually be obtained from a combination of different approaches including behavioral studies. Neuropeptides localized in fibers in the LC
Localized in fibres
Authors CaffC et al., 1985 De Vries et al., 1985 Pickel et al., 1979 Watson et al., 1978 Cummings et al., 1983 Finley et al., 1981 Johansson et al., 1984 Everitt et al., 1984 Chronwall et al., 1985 Charney et al., 1982 Pickel et al., 1979 Jennes et al., 1982 Moore and Gustafson, 1989 Feldman and Kastin, 1984
may be considered as potential transmitter candidates, but the question of whether the fibers make direct synaptic contacts with NA cell bodies or dendrites is not yet known. It is also unclear whether the peptides are colocalized with any one of the two major afferent fiber systems projecting to this brain area, as described by AstonJones et al., 1986 (see also this volume). With such information available, the electrophysiological experiments could be designed in a more appropriate manner. The knowledge of the precise termination area of these fibers would point to the sites where peptides are likely to exert their actions i.e., in the cell body area of the LC proper or on their dendrites. Neuropeptides colocalized with NA LC neurons, on the other hand, may be cotransmitter candidates, i.e., they may be transported to target areas and released as cotransmitters together with noradrenaline. Among the neuropeptides shown to be colocalized with noradrenaline are galanin (Melander et al., 1986), neuropeptide Y (Everitt et al., 19841, enkephalin (Charnay et al., 1982) vasopressin (Caff6 et al., 1985) and the corticotropin-releasing factor (Cummings et al., 1983). Finally, a release of peptides via dentrites or axon collaterals located within the LC area itself is also conceivable. The
243
TABLE 2 Electrophysiological effects of neuropeptides on locus coeruleus neurons Preparation
Authors
increase increase increase
anaesthetized rat anaesthetized rat slice, gerbil
Olpe and Baltzer, 1981 Berecek eta/., 1987 Olpe et al., 1987
substance P
increase increase increase
anaesthetized rat slice, rat slice, gerbil
Guyenet and Aghajanian, 1977 Cheeseman et al., 1983 Olpe et al., 1987
adrenocorticotropin hormone, ACTH
increase variable effects increase
anaesthetized rat anaesthetized rat slice, gerbil
Olpe and Jones, 1982 A d a m and Foot, 1988 Olpe et al., 1987
corticotropin-releasing factor, CRF
increase
anaesthetized rat
Valentino et al., 1983
somatostatin, SRlF
decrease
slice, gerbil cell culture
Olpe ef al., 1987 Masuko et al., 1986
neuropeptide Y
decrease
slice, rat
Illes and Regenold, 1990
enkephalin
decrease
anaesthetized rat slice, rat
Korf et al., 1974 Williams and North, 1984 Pepper and Henderson 1980
neurotensin
inactive decrease increase
anaesthetized rat anaesthetized rat slice, gerbil
Guyenet and Aghajanian, 1977 Young et al., 1978 Olpe et al., 1987
galanin
decrease
slice, rat
Seutin et al., 1989
Peptide
Effect on dicharge rate
vasopressin
Effect on membrane potential
hyperpolarisation hyperpolarisation
hyperpolarisation
peptides could thereby function to regulate LC neuronal activity at the level of NA cell bodies or dendrites. The major methodological problems linked to electrophysiological studies on peptides concern the mode of peptide administration, the peptide catabolism within the tissue and the diffusional barriers imposed on these rather large molecules by the tissue itself. Most studies published on the action of peptides in this area are complicated by these factors, with the exception possibly of cellculture studies (Masuko et al., 1986). The route of administration may be of particular importance. The mode of application of somatostatin in the hippocampus has been shown to give rather variable results with local ionophoretic administrations resulting in excitatory (Olpe et aL, 1980), and bath-applications exerting inhibitory effects
(Pittman and Siggins, 1981). Another major problem of this kind of investigation is the fact that receptor blockers of these peptides were either not available or were not used. In view of these methodological considerations and limitations, and given the fact that the cellular origin of most peptides is unknown, our understanding of the action of neuropeptides in the LC must be considered as rather preliminary. The effects of peptides on locus coeruleus neurons
Table 2 lists some of the major effects described with neuropeptides on rodent LC neurons. With the exception of a few investigations, we lack detailed information on these agents as modulators of established afferent transmitters or of
245 E ffect on f irin g rate of locus coeruleus neurons
-
490 -
N=10 7
Neurok,inin - A Neurokinin - B Substance - P Eledoisin
.-C u
ol C
Concentratlon in nM
Fig. 2. Mean concentration-response curves of the excitatory actions of four tachykinins on the spontaneous firing rate of gerbil LC neurons are depicted. The spontaneous mean firing rate recorded 10 min prior to the peptide administration was taken as the control value. Each value is the mean of 10 neurons rt S.E.M.
neurotensin was found to be inactive in viuo (Guyenet and Aghajanian, 1977) and subsequently was observed to depress NA cells in a similar study (Young et al., 1978). Bath-applied neurotensin in the slice preparation of the gerbil was weakly excitatory if tested at a concentration of 10 p M (Olpe et al., 1987). Taken together, a rather small number of neurons has been investi.34
SUBSTANCE P
NEUROKlNlN A
. 34
ELEDOlSlN
NEUROKlNlN B
Fig. 3. The effect of prolonged bath-administration of four tachykinins on the spontaneous firing rate of four different LC neurons of the gerbil is depicted. Each peptide was administered at a concentration of 100 WM. A quite pronounced tachyphylaxis is seen with substance P.
gated so far and the cellular sensitivity may not be uniform across the LC. Pronounced excitation of neuronal activity was observed however with the vasoactive intestinal peptide in an in vitro study (Wang and Aghajanian, 1989). It was concluded that this peptide induces an Na+-dependent inward current and the involvement of a pertussis toxin-sensitive G protein was suggested (Wang and Aghajanian, 1989). Considerable immunoreactivity to angiotensin I1 has been observed recently in the LC area (see the chapters by Marshall et al. and Speth et al.). The binding site for angiotensin was described as predominantly of the A 11, type (see Speth et al., this volume). It was found that angiotensin 11, while having almost no effect on the spontaneous discharge frequency or on the membrane potential of LC neurons, appears to attenuate glutamate-evoked excitatory responses (see Marshall et al., this volume). Both agents were pressure ejected. In three experiments, we tested the action of bath-applied angiotensin I1 on eight rat LC neurons. The peptide was applied at concentrations of 10 and 100 p M . It did not induce any notable change in spontaneous cell firing (OIpe, unpublished observation). Taken together the results suggest that angiotensin I1 may act as a neuromodulator in the LC area. A modulatory role has also been proposed for enkephalin (McFadzean et al., 1987) and recently for neuropeptide Y (Illes and Regenold, 1990). Immunoreactivity to dynorphin, a putative ligand for kappa opioid receptors, has been localized in the LC area (Zamir et al., 1983). On the basis of electrophysiological investigations performed on a LC preparation, it has been suggested that K receptors are located presynaptically on afferent fibers that provide an excitatory input to LC. Activation of K receptors by a selective agonist attenuated locally evoked excitatory postsynaptic potentials (McFadzean et al., 1987). Neuropeptide Y exerts two effects. It depresses LC neuronal activity (Illes and Regenold, 1990) and potentiates the hyperpolarizing effect of a,-receptor agonists in a selective manner
245 E ffect on f irin g rate of locus coeruleus neurons
-
490 -
N=10 7
Neurok,inin - A Neurokinin - B Substance - P Eledoisin
.-C u
ol C
Concentratlon in nM
Fig. 2. Mean concentration-response curves of the excitatory actions of four tachykinins on the spontaneous firing rate of gerbil LC neurons are depicted. The spontaneous mean firing rate recorded 10 min prior to the peptide administration was taken as the control value. Each value is the mean of 10 neurons rt S.E.M.
neurotensin was found to be inactive in viuo (Guyenet and Aghajanian, 1977) and subsequently was observed to depress NA cells in a similar study (Young et al., 1978). Bath-applied neurotensin in the slice preparation of the gerbil was weakly excitatory if tested at a concentration of 10 p M (Olpe et al., 1987). Taken together, a rather small number of neurons has been investi.34
SUBSTANCE P
NEUROKlNlN A
. 34
ELEDOlSlN
NEUROKlNlN B
Fig. 3. The effect of prolonged bath-administration of four tachykinins on the spontaneous firing rate of four different LC neurons of the gerbil is depicted. Each peptide was administered at a concentration of 100 WM. A quite pronounced tachyphylaxis is seen with substance P.
gated so far and the cellular sensitivity may not be uniform across the LC. Pronounced excitation of neuronal activity was observed however with the vasoactive intestinal peptide in an in vitro study (Wang and Aghajanian, 1989). It was concluded that this peptide induces an Na+-dependent inward current and the involvement of a pertussis toxin-sensitive G protein was suggested (Wang and Aghajanian, 1989). Considerable immunoreactivity to angiotensin I1 has been observed recently in the LC area (see the chapters by Marshall et al. and Speth et al.). The binding site for angiotensin was described as predominantly of the A 11, type (see Speth et al., this volume). It was found that angiotensin 11, while having almost no effect on the spontaneous discharge frequency or on the membrane potential of LC neurons, appears to attenuate glutamate-evoked excitatory responses (see Marshall et al., this volume). Both agents were pressure ejected. In three experiments, we tested the action of bath-applied angiotensin I1 on eight rat LC neurons. The peptide was applied at concentrations of 10 and 100 p M . It did not induce any notable change in spontaneous cell firing (OIpe, unpublished observation). Taken together the results suggest that angiotensin I1 may act as a neuromodulator in the LC area. A modulatory role has also been proposed for enkephalin (McFadzean et al., 1987) and recently for neuropeptide Y (Illes and Regenold, 1990). Immunoreactivity to dynorphin, a putative ligand for kappa opioid receptors, has been localized in the LC area (Zamir et al., 1983). On the basis of electrophysiological investigations performed on a LC preparation, it has been suggested that K receptors are located presynaptically on afferent fibers that provide an excitatory input to LC. Activation of K receptors by a selective agonist attenuated locally evoked excitatory postsynaptic potentials (McFadzean et al., 1987). Neuropeptide Y exerts two effects. It depresses LC neuronal activity (Illes and Regenold, 1990) and potentiates the hyperpolarizing effect of a,-receptor agonists in a selective manner
246
(Illes and Regenold, 1990). The site and mechanism of this interaction remain to be elucidated. Discussion
The available data on neuropeptides in the LC of the rodent brain suggests that they play a physiological role in regulating neuronal activity. This hypothesis remains to be confirmed, however. There are no studies yet showing that blockers of any one of these transmitter or cotransmitter candidates affect LC neuronal activity. More importantly, it would be crucial to study the effect of stimulating the peptidergic neurons which provide fibers to the LC to see if the synaptic effects mimic the action of local or bath-applied peptides. In view of the lack of such information, our knowledge on the action and role of peptides in the LC is fragmentary and preliminary. So far, the strongest evidence for a role of various neuropeptides in this region is the histochemical demonstration that they are present. It is clear that unless we advance in our understanding of their functions, we will lack important information on the physiology of LC neurons. It was noted above that the apparent discrepancy between the homogeneity of the LC and the rather large number of postulated functions poses one of the main problems remaining to be solved. So far, the work related to peptides does not help much in approaching this issue. However, the observation that some peptides are not homogeneously distributed could be indicative of subgroups of neurons which might have different functions. Subpopulations of LC perikarya have been shown to contain neuropeptide Y and galanin (Moore and Gustafson, 1989). Coexistence of vasopressin with noradrenaline has been demonstrated mainly in the posterior part of LC (Caffk et al., 1985). In a recent study on the distribution of substance P, neuropeptide Y, neurotensin and thyrotropin-releasing hormone in the human LC it was demonstrated that these peptides are unevenly distributed (Pammer et al., 1990). The
immunoreactive neuronal networks showed the highest density in the middle, and to a lesser extent, in the caudal part of this nucleus. It is thus conceivable that the study of neuropeptides might provide a means of dividing this small nucleus into compartments of functional neuronal subgroups. It is not unlikely that these peptides will be shown to have important functions in the control of the LC and that the knowledge of their role might change our view on the functions of the LC itself. Acknowledgement
The author thanks Mrs. M. Koller for typing the manuscript. References Adams, L.M. and Foote, S.L. (1988) Effects of locally infused pharmacological agents on spontaneous and sensoryevoked activity of locus coeruleus neurons. Bruin Res. Bull., 21: 395-400. Aston-Jones, G., Ennis M., Pieribone, V.A., Nickell, W.T. and Shipley, M.T. (1986) The brain nucleus locus coeruleus: Restricted afferent control of a broad efferent network. Science, 234: 734-737. Berecek, K.H., Olpe, H.-R., Mah, S.C. and Hofbauer, K.G. (1987) Alterations in responsiveness of noradrenergic neurons of the locus coeruleus in deoxycorticosterone acetate (DOCAI-salt hypertensive rats. Brain Res., 401: 303-31 1. Cafft, A.R., Van Leeuwen, F.W., Buijs, R.M., De Vries, G.J. and Geffard, M. (1985) Coexistence of vasopressin, neurophysin and noradrenaline in medium-sized cells of the locus coeruleus and subcoeruleus in the rat. Brain Res., 338: 160-164. Charnay, Y., LCger, L., Dray, F., Btrod, A,, Jouvet, M., Pujol, J.E. and Dubois, P.M. (1982) Evidence for the presence of enkephalin in catecholaminergic neurons of cat locus coeruleus. Neurosci. Lett., 30: 147-151. Cheeseman, H.J., Pinnock, R.D. and Henderson, G. (1983) Substance P excitation of rat locus coeruleus neurons. Eur. J. Pharmacol., 94: 93-99. Chronwall, B.M., Di Maggio, D.A., Massari, V.J., Pickel, V.M., Ruggiero, D.A. and O’Donohue, T.L. (1985) The anatomy of neuropeptide-Y-containing neurons in rat brain. Neuroscience, 15: 1159-1181. Cummings, S., Elde, R., Ells, J. and Lindall, A. (1983) Corticotropin-releasing factor immunoreactivity is widely distributed within the central nervous system of the rat: An immunohistochemical study. J. Neurosci., 3: 135551368,
247 De Vries, G.J., Buijs, R.M., Van Leeuwen, F.W., Caff6, A.R. and Swabb, D.F. (1985) The vasopressinergic innervation of the brain in normal and castrated rats. J. Comp. Neurol., 233: 236-254. Douglas, F.L., Palkovits, M. and Brownstein, M.J. (1982) Regional distribution of substance P-like immunoreactivity in the lower brainstem of the rat. Brain Res., 245: 376-378. Everitt, B.J., Hokfelt, T., Terenius, R., Tatemoto, K., Mutt, V. aild Goldstein, M. (1984) Differential co-existence of neuropeptide Y (NPY)-like immunoreactivity with catecholamines in the central nervous system of the rat. Neuroscience, 11: 443-462. Feldman, S.C. and Kastin, A.J. (1984) Localization of neurons containing immunoreactive delta sleep-inducing peptide in the rat brain: An immunocytochemical study. Neuroscience, 11: 443-462. Finley, J.C.W., Maderdrut, J.L., Roger, L.J. and Petrusz, P. (1981) The immunocytochemical localization of somatostatin-containing neurons in the rat central nervous system. Neuroscience, 6: 2173-2192. Grzanna, R., Chee, W.K. and Akeyson, E.W. (1987) Noradrenergic projections to brainstem nuclei: Evidence for differential projections from noradrenergic subgroups. J. Comp. Neurol., 263: 76-91. Guyenet, P.G. and Aghajanian, G.K. (1977) Excitation of neurons in the nucleus locus coeruleus by substance P and related peptides. Brain Res., 136: 178-188. Illes, P. and Regenold, J.T. (1990) Interaction between neuropeptide Y and noradrenaline on central catecholamine neurons. Nature (London), 344: 62-63. Jennes, L., Stumpf, W.E. and Kalivas, P.W. (1982) Neurotensin: Topographical distribution in rat brain by immunohistochemistry. J. Comp. Neurol., 210: 21 1-224. Johansson, O., Hokfelt, T. and Elde, R.P. (1984) Immunocytochemical distribution of somatostatin-like immunoreactivity in the central nervous system of the adult rat. Neuroscience, 13: 265-339. Korf, J., Bunney, B.S. and Aghajanian, G.K. (1974) Noradrenergic neurons: Morphine inhibition of spontaneous activity. Eur. J. Pharmacol., 25: 165-175. Masuko, S., Nakajima, Y., Nakajima, S. and Yamaguchi, K. (1986) Noradrenergic neurons from the locus coeruleus in dissociated cell culture: Culture methods, morphology and electrophysiology. J. Neurosci., 6: 3229-3241. McFadzean, I., Lacey, M.G., Hill, R.G. and Henderson, G. (1987) Kappa opioid receptor activation depresses excitatory synaptic input to rat locus coeruleus neurons in vitro. Neuroscience, 20: 231-239. Melander, T., Hokfelt, T., Rokaeus, A,, Cuello, A.C., Oertel, W.H., Verhofstad, A. and Goldstein, M. (1986) Coexistence of galanin-like immunoreactivity with catecholamines, 5-hydroxytryptamine, GABA and neuropeptides in the rat CNS. J. Neurosci., 6: 3640-3654. Moore, R.Y. and Gustafson, E.L. (1989) The distribution of dopamine-p-hydroxylase, neuropeptide Y and galanin in locus coeruleus neurons. J. Chem. Neuroanat., 2: 95-106. Olpe, H.-R. and Baltzer, V. (1981) Vasopressin activates
noradrenergic neurons in the rat locus coeruleus: A microionophoretic investigation. Eur. J. Pharmacol., 73: 377378. Olpe, H.-R. and Jones, R.S.G. (1982) Excitatory effects of ACTH on noradrenergic neurons of the locus coeruleus in the rat. Brain Res., 251: 177-179. Olpe, H.-R., Balcar, V.J., Bittiger, H., Rink, H. and Sieber, P. (1980) Central actions of somatostatin. Eur. J. Pharmacol., 63: 127-133. Olpe, H.-R., Steinmann, M.W. and Jones, R.S.G. (1985) Electrophysiological perspectives on locus coeruleus: Its role in cognitive versus vegetative functions. Physiol. Psycho/., 13: 179-187. Olpe, H.-R., Steinmann, M.W., Pozza, M.F. and Haas, H.L. (1987) Comparative investigations of the actions of ACTH 1-24' somatostatin, neurotensin, substance P and vasopressin on locus coeruleus neuronal activity in vitro. Naunyn Schmiedebergs Arch. Pharmacol., 336: 434-437. Palkovits, M., Epelbaum, J., Tapia-Arancibia, L. and Kordon, C. (1982) Somatostatin in catecholamine-rich nuclei of the brainstem. Neuropeptides, 3: 139-144. Pammer, C., Gorcs, T. and Palkovits, M. (1990) Peptidergic innervation of the locus coeruleus cells in the human brain. Brain Res., 515: 247-255. Pepper, C.M. and Henderson, G. (1980) Opiates and opioid peptides hyperpolarize locus coeruleus neurons in vitro. Science, 209: 394-396. Pickel, V.M., John, T.H., Reis, D.J., Leeman, S.E. and Miller, R.J. (1979) Electron microscopic localization of substance P and enkephalin in axon terminals related to dendrites of catecholaminergic neurons, Brain Res., 160: 387-400. Pittmann, Q.R. and Siggins, G.R. (1981) Somatostatin hyperpolarizes hippocampal pyramidal cells in vitro. Brain Rex, 221: 402-408. Seutin, V., Verbanck, P., Masotte, L. and Dresse, A. (1989) Galanin decreases the activity of locus coeruleus neurons in vitro. Eur. J. Pharmacol., 164: 373-376. Svensson, T.H. (1987) Peripheral, autonomic regulation of locus coeruleus noradrenergic neurons in brain: Putative implications for psychiatry and psychopharmacology. Psychopharmacology, 92: 1-7. Valentino, R.J., Foote, S.L. and Aston-Jones G. (1983). Corticotropin-releasing factor activates noradrenergic neurons of the locus coeruleus. Brain Res., 270: 363-367. Wang, Y.-Y. and Aghajanian, G.K. (1989) Excitation of locus coeruleus neurons by vasoactive intestinal peptide: Evidence for a g-protein-mediated inward current. Brain Res., 500: 107-118. Watson, S.J., Richard, C.W. and Barchas, J.D. (1978) Adrenocorticotropin in rat brain: Immunocytochemical localization in cells and axons. Science, 20: 1180-1181. Williams, J.T. and North, R.A. (1984) Opiate-receptor interactions on single locus coeruleus neurons. Mol. PharmaC O ~ . 26: , 489-497. Williams, J.T., Egan, T.M. and North, R.A. (1982) Enkephalin opens potassium channels in mammalian central neurons. Nature (London), 299: 74-76.
248 Williams, J.T., North, R.A., Shefner, S.A., Nishi, S. and Egan, T.M. (1984) Membrane properties of rat locus coeruleus neurons. Neuroscience, 13: 137-156. Young, W.S., Uhl, G.R. and Kuhar, M.J. (1978) Ionophoresis of neurotensin in the area of the locus coeruleus. Brain Res.. 150: 431-435.
Zamir, N., Palkovits, M. and Brownstein, M.J. (1983) Distribution of immunoreactive dynorphin in the central nervous system of the rat. Bruin Rex, 280: 81-93.
24 1 CHAPTER 18
Responses of locus coeruleus neurons to neuropeptides H.-R. Olpe and M. Steinmann Research and Deaelopment Department, Pharmaceuticals Division, CIBA-GEIGY Ltd., Basel, Switzerland
The knowledge on the neuronal inputs to the locus coeruleus (LC) and their roles in regulating noradrenergic (NA) cellular activity is quite advanced. In recent years, however, about ten neuropeptides were found to be localized in the area of the rodent LC; peptides which may be considered as potential transmitters o r modulators acting in this area. Electrophysiological studies performed in viuo and in Liitro have revealed that many of these peptides are able to alter LC neuronal activity. Stimulatory effects have been described with vasopressin, substance P, adrenocorticotropin hormone and corticotropin-releasing factor. Depressant effects were seen with galanin, somatostatin, neuropeptide Y and enkephalin. Variable actions were observed in the case of neurotensin. While these findings point to a possible regulatory function of these peptides in this area, precise roles
remain unclear. Important information is lacking that would conclusively demonstrate their regulatory functions. It should be determined whether the stimulation of peptidergic cells elicits synaptic effects identical to the ones observed with local exogenous peptide applications. By studying the action of blockers of these transmitter and modulator candidates, we would probably begin to understand their importance in the regulation of tonic and phasic activity components. The LC is generally considered to consist of a homogenous group of neurons. T h e recent observation that subpopulations of these cells contain peptides as in the case of neuropeptide Y, galanin and vasopressin, points to the possible existence of subgroups of neurons having different functions.
Key words: locus coeruleus, neuropeptides, electrophysiology, modulator, rat
Introduction
The locus coeruleus (LC) of the rat brain is generally considered to be a homogenous and compact nucleus containing mainly noradrenergic (NA) neurons. Intra- and extracellular electrophysiological recordings are in keeping with this notion since the biophysical properties of all cells appear to be very similar (Williams et al., 1984). Over the past ten years, a steadily increasing number of potential physiological roles have been proposed that are claimed to be linked with this
brain area (Olpe et al., 1985; Svensson, 1987). We are, therefore, facing the problem that a small number of seemingly homogeneous neurons is thought to be involved in several distinct brain functions. One theoretical approach for solving this problem is to search for a common basic action of noradrenaline that would be coupled with a number of different, more complex functions. Alternatively, a basis for the heterogeneity of functions could be found in the fact that the LC projections have recently been shown to be selective and not as widely and diffusely spread
242
TABLE 1 Neuropeptides present in the area of the locus coeruleus Peptides present in locus coeruleus
Localized in perikarya
vasopressin substance P adrenocorticotropin hormone, ACTH corticotropin-releasing factor, CRF somatostatin. SRIF neuropeptide Y enkephalin neurotensin galanin delta sleep-inducing peptide, DSIP
as originally described (Grzanna et al., 1987). Another possible suggestion for the heterogeneity of LC neurons themselves derives from recent reports that several neuropeptides are present in this brain area which have been localized in fibers and/or cell bodies. A survey of the kind of peptides described in this area is provided in Table 1 (see also Sutin and Jacobowitz, this volume). It is remarkable that the rodent LC contains not only a considerable number of different peptides, but some of them are present at rather high concentrations, as in the case of substance P (Douglas et al., 1982) and somatostatin (Palkovits et al., 1982). The presence of these peptides raises the question of their physiological functions. Electrophysiological techniques provide a means of addressing this issue, at least partly, in that the sensitivity of LC neurons to these peptides can be investigated. While such studies give valuable information on the effects of peptides on cell activity, their membrane potentials and other biophysical membrane properties, they do not necessarily solve the problem of identifying the peptides’ physiological roles. Such information may eventually be obtained from a combination of different approaches including behavioral studies. Neuropeptides localized in fibers in the LC
Localized in fibres
Authors CaffC et al., 1985 De Vries et al., 1985 Pickel et al., 1979 Watson et al., 1978 Cummings et al., 1983 Finley et al., 1981 Johansson et al., 1984 Everitt et al., 1984 Chronwall et al., 1985 Charney et al., 1982 Pickel et al., 1979 Jennes et al., 1982 Moore and Gustafson, 1989 Feldman and Kastin, 1984
may be considered as potential transmitter candidates, but the question of whether the fibers make direct synaptic contacts with NA cell bodies or dendrites is not yet known. It is also unclear whether the peptides are colocalized with any one of the two major afferent fiber systems projecting to this brain area, as described by AstonJones et al., 1986 (see also this volume). With such information available, the electrophysiological experiments could be designed in a more appropriate manner. The knowledge of the precise termination area of these fibers would point to the sites where peptides are likely to exert their actions i.e., in the cell body area of the LC proper or on their dendrites. Neuropeptides colocalized with NA LC neurons, on the other hand, may be cotransmitter candidates, i.e., they may be transported to target areas and released as cotransmitters together with noradrenaline. Among the neuropeptides shown to be colocalized with noradrenaline are galanin (Melander et al., 1986), neuropeptide Y (Everitt et al., 19841, enkephalin (Charnay et al., 1982) vasopressin (Caff6 et al., 1985) and the corticotropin-releasing factor (Cummings et al., 1983). Finally, a release of peptides via dentrites or axon collaterals located within the LC area itself is also conceivable. The
243
TABLE 2 Electrophysiological effects of neuropeptides on locus coeruleus neurons Preparation
Authors
increase increase increase
anaesthetized rat anaesthetized rat slice, gerbil
Olpe and Baltzer, 1981 Berecek eta/., 1987 Olpe et al., 1987
substance P
increase increase increase
anaesthetized rat slice, rat slice, gerbil
Guyenet and Aghajanian, 1977 Cheeseman et al., 1983 Olpe et al., 1987
adrenocorticotropin hormone, ACTH
increase variable effects increase
anaesthetized rat anaesthetized rat slice, gerbil
Olpe and Jones, 1982 A d a m and Foot, 1988 Olpe et al., 1987
corticotropin-releasing factor, CRF
increase
anaesthetized rat
Valentino et al., 1983
somatostatin, SRlF
decrease
slice, gerbil cell culture
Olpe ef al., 1987 Masuko et al., 1986
neuropeptide Y
decrease
slice, rat
Illes and Regenold, 1990
enkephalin
decrease
anaesthetized rat slice, rat
Korf et al., 1974 Williams and North, 1984 Pepper and Henderson 1980
neurotensin
inactive decrease increase
anaesthetized rat anaesthetized rat slice, gerbil
Guyenet and Aghajanian, 1977 Young et al., 1978 Olpe et al., 1987
galanin
decrease
slice, rat
Seutin et al., 1989
Peptide
Effect on dicharge rate
vasopressin
Effect on membrane potential
hyperpolarisation hyperpolarisation
hyperpolarisation
peptides could thereby function to regulate LC neuronal activity at the level of NA cell bodies or dendrites. The major methodological problems linked to electrophysiological studies on peptides concern the mode of peptide administration, the peptide catabolism within the tissue and the diffusional barriers imposed on these rather large molecules by the tissue itself. Most studies published on the action of peptides in this area are complicated by these factors, with the exception possibly of cellculture studies (Masuko et al., 1986). The route of administration may be of particular importance. The mode of application of somatostatin in the hippocampus has been shown to give rather variable results with local ionophoretic administrations resulting in excitatory (Olpe et aL, 1980), and bath-applications exerting inhibitory effects
(Pittman and Siggins, 1981). Another major problem of this kind of investigation is the fact that receptor blockers of these peptides were either not available or were not used. In view of these methodological considerations and limitations, and given the fact that the cellular origin of most peptides is unknown, our understanding of the action of neuropeptides in the LC must be considered as rather preliminary. The effects of peptides on locus coeruleus neurons
Table 2 lists some of the major effects described with neuropeptides on rodent LC neurons. With the exception of a few investigations, we lack detailed information on these agents as modulators of established afferent transmitters or of
245 E ffect on f irin g rate of locus coeruleus neurons
-
490 -
N=10 7
Neurok,inin - A Neurokinin - B Substance - P Eledoisin
.-C u
ol C
Concentratlon in nM
Fig. 2. Mean concentration-response curves of the excitatory actions of four tachykinins on the spontaneous firing rate of gerbil LC neurons are depicted. The spontaneous mean firing rate recorded 10 min prior to the peptide administration was taken as the control value. Each value is the mean of 10 neurons rt S.E.M.
neurotensin was found to be inactive in viuo (Guyenet and Aghajanian, 1977) and subsequently was observed to depress NA cells in a similar study (Young et al., 1978). Bath-applied neurotensin in the slice preparation of the gerbil was weakly excitatory if tested at a concentration of 10 p M (Olpe et al., 1987). Taken together, a rather small number of neurons has been investi.34
SUBSTANCE P
NEUROKlNlN A
. 34
ELEDOlSlN
NEUROKlNlN B
Fig. 3. The effect of prolonged bath-administration of four tachykinins on the spontaneous firing rate of four different LC neurons of the gerbil is depicted. Each peptide was administered at a concentration of 100 WM. A quite pronounced tachyphylaxis is seen with substance P.
gated so far and the cellular sensitivity may not be uniform across the LC. Pronounced excitation of neuronal activity was observed however with the vasoactive intestinal peptide in an in vitro study (Wang and Aghajanian, 1989). It was concluded that this peptide induces an Na+-dependent inward current and the involvement of a pertussis toxin-sensitive G protein was suggested (Wang and Aghajanian, 1989). Considerable immunoreactivity to angiotensin I1 has been observed recently in the LC area (see the chapters by Marshall et al. and Speth et al.). The binding site for angiotensin was described as predominantly of the A 11, type (see Speth et al., this volume). It was found that angiotensin 11, while having almost no effect on the spontaneous discharge frequency or on the membrane potential of LC neurons, appears to attenuate glutamate-evoked excitatory responses (see Marshall et al., this volume). Both agents were pressure ejected. In three experiments, we tested the action of bath-applied angiotensin I1 on eight rat LC neurons. The peptide was applied at concentrations of 10 and 100 p M . It did not induce any notable change in spontaneous cell firing (OIpe, unpublished observation). Taken together the results suggest that angiotensin I1 may act as a neuromodulator in the LC area. A modulatory role has also been proposed for enkephalin (McFadzean et al., 1987) and recently for neuropeptide Y (Illes and Regenold, 1990). Immunoreactivity to dynorphin, a putative ligand for kappa opioid receptors, has been localized in the LC area (Zamir et al., 1983). On the basis of electrophysiological investigations performed on a LC preparation, it has been suggested that K receptors are located presynaptically on afferent fibers that provide an excitatory input to LC. Activation of K receptors by a selective agonist attenuated locally evoked excitatory postsynaptic potentials (McFadzean et al., 1987). Neuropeptide Y exerts two effects. It depresses LC neuronal activity (Illes and Regenold, 1990) and potentiates the hyperpolarizing effect of a,-receptor agonists in a selective manner
245 E ffect on f irin g rate of locus coeruleus neurons
-
490 -
N=10 7
Neurok,inin - A Neurokinin - B Substance - P Eledoisin
.-C u
ol C
Concentratlon in nM
Fig. 2. Mean concentration-response curves of the excitatory actions of four tachykinins on the spontaneous firing rate of gerbil LC neurons are depicted. The spontaneous mean firing rate recorded 10 min prior to the peptide administration was taken as the control value. Each value is the mean of 10 neurons rt S.E.M.
neurotensin was found to be inactive in viuo (Guyenet and Aghajanian, 1977) and subsequently was observed to depress NA cells in a similar study (Young et al., 1978). Bath-applied neurotensin in the slice preparation of the gerbil was weakly excitatory if tested at a concentration of 10 p M (Olpe et al., 1987). Taken together, a rather small number of neurons has been investi.34
SUBSTANCE P
NEUROKlNlN A
. 34
ELEDOlSlN
NEUROKlNlN B
Fig. 3. The effect of prolonged bath-administration of four tachykinins on the spontaneous firing rate of four different LC neurons of the gerbil is depicted. Each peptide was administered at a concentration of 100 WM. A quite pronounced tachyphylaxis is seen with substance P.
gated so far and the cellular sensitivity may not be uniform across the LC. Pronounced excitation of neuronal activity was observed however with the vasoactive intestinal peptide in an in vitro study (Wang and Aghajanian, 1989). It was concluded that this peptide induces an Na+-dependent inward current and the involvement of a pertussis toxin-sensitive G protein was suggested (Wang and Aghajanian, 1989). Considerable immunoreactivity to angiotensin I1 has been observed recently in the LC area (see the chapters by Marshall et al. and Speth et al.). The binding site for angiotensin was described as predominantly of the A 11, type (see Speth et al., this volume). It was found that angiotensin 11, while having almost no effect on the spontaneous discharge frequency or on the membrane potential of LC neurons, appears to attenuate glutamate-evoked excitatory responses (see Marshall et al., this volume). Both agents were pressure ejected. In three experiments, we tested the action of bath-applied angiotensin I1 on eight rat LC neurons. The peptide was applied at concentrations of 10 and 100 p M . It did not induce any notable change in spontaneous cell firing (OIpe, unpublished observation). Taken together the results suggest that angiotensin I1 may act as a neuromodulator in the LC area. A modulatory role has also been proposed for enkephalin (McFadzean et al., 1987) and recently for neuropeptide Y (Illes and Regenold, 1990). Immunoreactivity to dynorphin, a putative ligand for kappa opioid receptors, has been localized in the LC area (Zamir et al., 1983). On the basis of electrophysiological investigations performed on a LC preparation, it has been suggested that K receptors are located presynaptically on afferent fibers that provide an excitatory input to LC. Activation of K receptors by a selective agonist attenuated locally evoked excitatory postsynaptic potentials (McFadzean et al., 1987). Neuropeptide Y exerts two effects. It depresses LC neuronal activity (Illes and Regenold, 1990) and potentiates the hyperpolarizing effect of a,-receptor agonists in a selective manner
246
(Illes and Regenold, 1990). The site and mechanism of this interaction remain to be elucidated. Discussion
The available data on neuropeptides in the LC of the rodent brain suggests that they play a physiological role in regulating neuronal activity. This hypothesis remains to be confirmed, however. There are no studies yet showing that blockers of any one of these transmitter or cotransmitter candidates affect LC neuronal activity. More importantly, it would be crucial to study the effect of stimulating the peptidergic neurons which provide fibers to the LC to see if the synaptic effects mimic the action of local or bath-applied peptides. In view of the lack of such information, our knowledge on the action and role of peptides in the LC is fragmentary and preliminary. So far, the strongest evidence for a role of various neuropeptides in this region is the histochemical demonstration that they are present. It is clear that unless we advance in our understanding of their functions, we will lack important information on the physiology of LC neurons. It was noted above that the apparent discrepancy between the homogeneity of the LC and the rather large number of postulated functions poses one of the main problems remaining to be solved. So far, the work related to peptides does not help much in approaching this issue. However, the observation that some peptides are not homogeneously distributed could be indicative of subgroups of neurons which might have different functions. Subpopulations of LC perikarya have been shown to contain neuropeptide Y and galanin (Moore and Gustafson, 1989). Coexistence of vasopressin with noradrenaline has been demonstrated mainly in the posterior part of LC (Caffk et al., 1985). In a recent study on the distribution of substance P, neuropeptide Y, neurotensin and thyrotropin-releasing hormone in the human LC it was demonstrated that these peptides are unevenly distributed (Pammer et al., 1990). The
immunoreactive neuronal networks showed the highest density in the middle, and to a lesser extent, in the caudal part of this nucleus. It is thus conceivable that the study of neuropeptides might provide a means of dividing this small nucleus into compartments of functional neuronal subgroups. It is not unlikely that these peptides will be shown to have important functions in the control of the LC and that the knowledge of their role might change our view on the functions of the LC itself. Acknowledgement
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