- Email: [email protected]
HYPOTHALAMIC PEPTIDES: NEW EVIDENCE FOR " PEPTIDERGIC " PATHWAYS IN THE C.N.S.
393
previous report 240 in which the P.C.A. extracts of colonic
Hypothesis
carcinomas could be separated into C.E.A.-active and i-active fractions. And it is likely that the incomplete
blood-group-like activity which was previously reported in a C.E.A. preparation 12 was caused by the presence of contaminant precursor substances. Earlier studies of the carbohydrate and aminoacid compositions of the fractions a to e of the tumour 1 C.E.A. preparation had suggested that fraction a was contaminated with a mucin-type glycoprotein.18 It was not surprising, therefore, that this fraction had the
highest blood-group precursor-like activity. The biological role of the blood-group antigens is not yet understood; nor is it known whether their deficient synthesis in entodermal tumours and in their metastases is a manifestation of rapid tissue growth, or However, the present studies indicate that the precursorlike activities of these tumours deserve detailed investigation, as does the possible release of these antigens into the bloodstream. Sensitive radioimmunoassays are currently being designed for the accurate measurement of these antigens in tumours and their fractions. Thus, high titre anti-I and anti-i cold. agglutinins which recognise several types of determinants on blood-group precursors may well prove to be powerful reagents in the detection and monitoring of a "new" set of tumour-associated antigens.
in
some
way related to tumour invasiveness.
We thank Dr M. A. Bukhari for the monosaccharide analyses; Mr S. Patel for the radioimmunoassays; and Mr R. Childs and Miss J. W. Summers for technical assistance. Requests for reprints should be addressed to T. F. REFERENCES
1. Hakomori, S., Kijimoto, S., Siddiqui, B. in Membrane Research (edited by C. F. Fox); p. 253. New York, 1972. 2. Masamune, H., Kawasaki, H., Abe, S., Oyama, K., Yamaguchi, Y. Tohoku J. exp. Med. 1958, 68, 81. 3. Iseki, S., Furukawa, K., Ishikara, K. Proc. Jap. Acad. 1962, 38, 556. 4. Hakomori, S., Koscielak, J., Bloch, H., Jeanloz, R. J. Immun. 1967, 98, 31. 5. Davidsohn, I., Ni, L. Y., Stejskal, R. Archs Path. 1971, 92, 456. 6. Gold, R., Freedman, S. O. J. exp. Med. 1965, 121, 439. 7. Gold, R., Freedman, S. O. ibid. 1965, 122, 467. 8. Lo Gerfo, P., Krupey, J., Hansen, H. J. New Engl. J. Med. 1971, 285, 138. 9. Zamchek, N., Moore, T. L., Dhar, P., Kupchik, H. Z. ibid. 1972, 286, 83. 10. Martin, F., Martin, M. S. Int. J. Cancer, 1970, 6, 352. 11. Mach, J. P., Pusztaszeri, G. Immunochemistry, 1972, 9, 1031. 12. Simmons, D. A. R., Perlmann, P. Cancer Res. 1973, 33, 313. 13. Feizi, T., Kabat, E. A., Vicari, G., Anderson, B., Marsh, W. L. J. exp. Med. 1971, 133, 39. 14. Feizi, T., Kabat, E. A., Vicari, G., Anderson, B., Marsh, W. L. J. Immun. 1971, 106, 1578. 15. Feizi, T., Kabat, E. A. J. exp. Med. 1972, 135, 1247. 16. Krupey, J., Gold, P., Freedman, S. O. ibid. 1968, 128, 387. 17. Turberville, C., Darcy, D. A., Laurence, D. J. R., Johns, E. W., Neville, A. M. Immunochemistry, 1973, 10, 841. 18. Turberville, C. Unpublished. 19. Clamp, J. R., Bhatti, T., Chambers, R. E. Meth. biochem. Analysis, 1971, 19, 229. 20. Feizi, T., Cederqvist, L. L., Childs, R. Br. J. Hœmat. 1975, 30, 485. 21. Vicari, G., Kabat, E. A. J. Immun. 1969, 102, 821. 22. Vicari, G., Kabat, E. A. Biochemistry, 1970, 9, 3414. 23. Laurence, D. J. R., Stevens, U., Bettelheim, R., Darcy, D., Leese, C., Turberville, C., Alexander, P., Johns, E. W., Neville, A. M. Br. med. J. 1972, iii, 605. 24. Cooper, A. G., Brown, M. C., Kirsh, M. E., Rule, A. H. J. Immun.
1974, 113, 1246.
Addendum T antigen (a presumed precursor of blood-group MN antigens) has recently been reported in human breast cancers (Springer, G. F., Desai, P. R., Banatwala, I. y. natn. Cancer Inst. 1975, S4, 335).
HYPOTHALAMIC PEPTIDES: NEW EVIDENCE FOR " PEPTIDERGIC " PATHWAYS IN THE C.N.S.
JOSEPH B.
MARTIN
LEO P. RENAUD
PAUL BRAZEAU
Division of Neurology, Department of Montreal General Hospital and McGill Montreal, Canada
Sum ary
Medicine, University,
Hypothalamic peptides, discovered be-
cause of their effects on stimulation or inhibition of anterior-pituitary-hormone release, have now been shown to be distributed in many regions of the central nervous system (C.N.S.). Demonstration of their localisation in nerve terminals, of behavioural effects exclusive of anterior pituitary actions, and of depressant effects in electrophysiological experiments suggests a role in central-nervous-system function. It is postulated that a system of peptidergic neurons exists within the C.N.S. with axon terminations both on the pituitary portal system and on other neurons. THE neural control of endocrine homoeostasis has been extensively investigated. The isolation and structural characterisation of several hypothalamic peptides that are known to regulate anterior-pituitary-hormone secretion has led to the emergence of evidence that points to new concepts of brain-hormone interaction, and permits speculation with respect to the role of " neurosecretion " in central-nervous-system (c.N.s.) function. We review here some of the information on the role that peptides, particularly hypothalamic peptides, may have in the C.N.s. and speculate on the existence of a system of peptide neurosecretory hypothalamic neurons that may terminate, not only on the pituitary portal capillary plexus, but also in extrahypothalamic regions of the brain. Several independent observations have drawn attention to the possibility that low-molecular-weight shortchain polypeptides have important direct effects on brain function. For example, angiotensin n, which promotes aldosterone secretion from the adrenal gland, may also be synthesised within the C.N.S.l Hypothalamic injection of angiotensin 11 causes animals in a normal state of water balance to drink, and its direct application to certain hypothalamic neurons increases their spike-discharge activity.2 Adrenocorticotrophin (A.C.T.H.) and smaller peptide fragments of A.C.T.H. that are devoid of corticotropic activity can influence the behaviour of laboratory animals 3 The c.N.s. actions of A.C.T.H. may explain certain of the behavioural alterations that accompany excessive A.C.T.H. secretion in Cushing’s disease. Low-molecularweight peptides have also been implicated in the regulation of sleepand memory.5 It is now possible to extend such observations to the recently identified hypothalamic peptides that regulate secretion of anterior-pituitary trophic hormones
(T.R.H.,
thyrotropin-releasing
hormone;
G.N.R.H.,
394
gonadotropin-releasing hormone; and somatostatin/ growth-hormone release-inhibiting hormone).6 It is believed that these peptides are synthesised by socalled releasing factor (peptidergic or tuberoinfundibular) neurosecretory neurons located in or near the medial basal hypothalamus. The peptides are thought to be secreted intermittently into the pituitary portal circulation from median-eminence response to a variety of stimuli.7
axon
terminals in
development of specific and sensitive radioimmunoassays for each of these hypothalamic peptides has permitted study of their distribution and subcellular localisation. T.R.H. is distributed widely in the medial basal hypothalamus,s whereas G.N.R.H. is localised primarily in the tuberal (arcuate) nuclei.9 Somat6statin The
is present in the arcuate and ventromedial nuclei and in the preoptic area, with lesser concentrations in other
regions of hypothalamus.
These differing patterns of distribution argue for a certain selectivity of neuronal localisation within individual hypothalamic nuclei. Immunohistochemical and biochemical studies of hypothalamic tissue have shown T.R.H.,l1 G.N.R.H.,’ and somatostatin 13,14 to be concentrated primarily within nerve terminals and synaptosomes. T.R.H. has been shown to be synthesised in vitro by hypothalamic tissue obtained from both mammalian 15 and subTissue from cerebral cortex mammalian species.16 in the rat is, however, incapable of synthesising T.R.H./5 suggesting that the localisation of synthetic mechanisms resides in the hypothalamus. The development of radioimmunoassays for the hypothalamic peptides has also resulted in the unexpected discovery that significant quantities of each peptide are present in extrahypothalamic regions, although concentrations are highest in the hypo-B thalamus. It is estimated in the rat that two-thirds of brain T.R.H. content is localised outside the hypothalamus 17; an extrahypothalamic localisation of T.R.H. has also been found in the human brain 18 G.N.R.H. is reported to be present in the cerebral cortex of the rat, and to show changes in concentration in response to stress.l9 Somatostatin is present in readily measureable quantities in the cerebral cortex, amygdala, spinal cord, and cerebrospinal fluid.2O We have shown that somatostatin within the cerebral cortex and amygdala is localised primarily within synaptosomes.21 Thus, it would appear that these peptides are located in nerve terminals in various regions of the brain. Administration of T.R.H., G.N.R.H., and somatostatin to laboratory animals and man results in behavioural effects not attributable directly to an action on the pituitary. T.R.H. has been suggested to have antidepressant properties in man,22 although there is controversy over its therapeutic efficacy.23.24 Probably related to a C.N.S. stimulatory role is the ability of T.R.H. to counteract the depressant effects of barbiturates, to antagonise ethanol narcosis, and to increase spontaneous motor activity in laboratory animals 2’ T.R.H. also reduces the potentially lethal effects of strychnine.28 G.N.R.H. restores copulatory behaviour in hypophyst.tomised rats,29 and appears to have aphrodisiac effects when administered systemically in man.3o Animal studies with somatostatin indicate that it has sedative effects and prolongs the activity of pentobarbital2s; it is also reported to potentiate the effects
of strychnine.28 Many of these effects remain after ablation (hypophysectomy, thyroidtarget-organ ectomy, or gonadectomy), indicating a significant direct That T.R.H. is present in the brain of C.N.S. action. primitive vertebrates that lack a pituitary or a thyroid 17 speaks eloquently for the primitive ancestry of the peptide and to the probability of a significant direct role in the C.N.S. Electrophysiological studies have provided evidence that polypeptides can influence neuronal responses. Microiontophoretic application of T.R.H., G.N.R.H., and somatostatin to single neurons is associated with a prominent but readily reversible decrease in excitability."-31 This effect is observed both in the hypothalamus and in several extrahypothalamic areas, including the cerebral cortex, cerebellurn, and spinal cord. The observation that only a certain population of neurons display peptide-sensitivity argues for a degree of selectivity and specificity of effect with respect to the individual peptides. If hypothalamic peptides are synthesised only by hypothalamic tissue, how do they become distributed Recent electrophysioto extrahypothalamic sites? logical studies have demonstrated evidence for axon collaterals of hypothalamic tuberoinfundibular neurons that terminate locally within the hypothalamus and also extend to brain sites quite distant from the hypothalamus-e.g., thalamic nuclei 34 Evidence of axon bifurcations of hypothalamic tuberoinfundibular neurons has been documented in anatomical studies.35 Significant hypothalamic projections to the amygdala,31 cerebral cortex,37 and brainstem 38 are known to exist. If diffusely distributed hypothalamic fibre systems can be shown to originate from neurons that produce the peptides (i.e., neurosecretory peptidergic neurons), this could explain the specific extrahypothalamic localisations of these substances. Reichlin and associates39 have reported that hypothalamic lesions in the thyroid regulatory area result in a reduction in T.R.H. content in the cerebral cortex. Taken together, these findings support the possi-
bility of a primary hypothalamic site for formation and distribution of these peptides. Activity in the tuberoinfundibular neurosecretory system could result in simultaneous release of peptides, not only into the capillary circulation in the median eminence, but also in other regions of the central nervous system that have synaptic connections from the hypothalamus. It is conceivable that release of these peptides from nerve terminals in extrahypothalamic regions of brain may subserve important biological functions such as feedback control and/or regulation of behaviour through a pre-synaptic or post-synaptic action affecting the electrical activity of distant C.N.S. neurons. In agreement with such an hypothesis for peptide modulation of neuronal excitability are the observations that all three hypothalamic peptides are now reported to be present in the pineal gland,2o,40 and that pinealectomy induces seizures in normal 41 and parathyroidectomised rats.42 Recent studies have implicated cyclic A.M.P. in central neuronal synaptic events 43 Since hypothalamic peptides act at a pituitary level by stimulating (T.R.H., G.N.R.H.) or inhibiting (somatostatin) cyclic A.M.P. formation,44. it is conceivable that this mechanism may
395 also contribute to their electrophysiological and behavioural effects in the C.N.S. The involvement of hypothalamic peptides in regulation of anterior-pituitary-hormone secretion may be a rather recent evolutionary development. This conclusion is supported by the relative lack of specificity of effect of the identified hypothalamic hormones on anterior pituitary. T.R.H. stimulates both T.S.H. and prolactin release 6; G.N.R.H. stimulates L.H. and F.S.H.44 Somatostatin in addition to inhibiting G.H. secretion also blocks T.R.H.-stimulated T.S.H. secretion and decreases insulin, glucagon, and gastrin release s The intermittent or episodic release of pituitary hormones, which is now recognised as the predominant form of hormone release." may be a reflection of underlying ongoing activity of basic C.N.S. rhythms. Homoeostasis in the periphery is accomplished by the prolonged metabolic effects of peripheral target hormones. Thus, stable hormonal control is achieved despite intermittent or rhythmic oscillations in hypothalamic neural function. We hypothesise, therefore, that hypothalamic peptidergic neurons, like monoaminergic neurons (dopaminergic, catecholaminergic, and serotoninergic) may be involved in the formation of a diffuse neural network that terminates in widespread regions of the C.N.S. The release of these peptides into the pituitary circulation may be but one facet of their biological significance. Their role in the C.N.S. may ultimately prove to be of equal or even greater importance in terms of neurobiological regulation. Proof of such a hypothesis will require further studies of the subcellular localisation of these peptides in extrahypothalamic tissue, demonstration of their release from non-hypothalamic nerve-endings after hypothalamic stimulation, and full characterisation of their putative role as either neurotransmitters or synaptic modulators. The intense interest in this question and the rapid emergence of data from several disciplines indicate that some answers will be forthcoming in the near future. Requests for reprints should be addressed to J. B. M., Division of Neurology, Montreal General Hospital, Suite 753, Livingston Hall, 1650 Cedar Avenue, Montreal, Quebe< H3G 1A4, Canada. REFERENCES 1. Reid, I. A., Ramsay, D. J., Malayan, S. A., Gänong, W. F. Program of the 57th Annual Meeting of Endocrine Society, June, 1975,
p. 138.
Fitzsimons, J. T. in Frontiers in Neuroendocrinology (edited by L. Martini and W. F. Ganong); p. 103. New York, 1971. 3. Greven, H. M., de Wied, D. Prog. Brain Res. 1973, 39, 429. 4. Pappenheimer, J. R., Fend, V., Koski, G. Proceedings of the 26th International Congress of Physiological Science, 1974, 11,
2.
abstr. 618. 5. Ungar, G. Biochem. Pharmac. 1974, 23, 1553. 6. Vale, W., Rivier, C. in Neuroendocrine Relationships (edited by L. Fisher). New York (in the press). 7. Martin, J. B., Renaud, L. P. ibid. 8. Brownstein, M., Palkovits, M., Saavedra, J. M., Bassiri, R. M., Utiger, R. D. Science, 1974, 185, 267. 9. Palkovits, M., Arimura, A., Brownstein, M., Schally, A. V., Saavedra, J. M. Endocrinology, 1974, 95, 554. 10. Brownstein, M., Arimura, A., Sato, H., Schally, A. V., Kizer, J. S. ibid. 1975, 96, 1456. 11. Barnea, A., Ben-Jonathan, N., Porter, J. C. Program of the 57th Annual Meeting of Endocrine Society, June, 1975, p. 95. 12. Zimmerman, E. A., Hsu, K. C., Ferin, M., Kozlowski, G. P. 13.
Endocrinology, 1974, 95, 1. Hokfelt, T., Efendic, S., Johansson, O., Luft, R., Arimura, Brain Res. 1974, 80, 165.
A.
Methods and Devices AN EXTRACTOR FOR SCALPEL BLADES
J. McKIE
A. SHAW
West of Scotland Health Boards, Department of Clinical Physics and Bio-Engineering,
Glasgow ALTHOUGH it is easy to attach disposable scalpel blades handles made to British or American standardstheir removal presents problems. Some of the smaller blades to
(e.g., Swann-Morton 10A, 11, 15) are especially difficult, and the task is not made easier when the hands are gloved and wet. There is a strong likelihood of a slipped tool or a snapped blade and a cut finger. At the Royal Infirmary, Glasgow, the injury most frequently reported was back injury sustained whilst lifting patients. Second place was shared by cuts received whilst discarding scalpel blades and scratches from used hypodermic needles when the proper disposal procedure had not been followed. We have devised a tool for extracting scalpel blades from their holders. The tool (fig. 1) has two jaws moved by " scissor-lever handles. The jaws are recessed to form a channel into which the scalpel handle extension fits, and the mating
14. Pelletier,
G., Labrie, F., Arimura, A., Schally, A. V. Am. Anat. J.
1974, 140, 445. 15. Reichlin, S., Mitnick, M. in Frontiers in Neuroendocrinology (edited by W. F. Ganong and L. Martini); p. 61. New York, 1973. 16. McKelvy, J. Brain Res. 1974, 65, 489. 17. Jackson, I. M. D., Reichlin, S. Endocrinology, 1974, 95, 854. 18. Winters, A. J., Eskay, R. L., Porter, J. C. J. clin. Endocr. Metab.
1974, 39, 960. 19. Montoya, E., Wilber, J., White, W., Gendrich, R. Program of the 57th Annual Meeting of Endocrine Society, June, 1975, p. 94. 20. Patel, Y. C., Weir, G. C., Reichlin, S. ibid. p. 127. 21. Tsang, D., Tan, S., Martin, J. B., Lal, S., Renaud, L. P., Brazeau, P. Program Society for Neuroscience, New York, November, 1975 (in the press). 22. Prange, A. J. (editor). The Thyroid Axis, Drugs and Behaviour. New York, 1974. 23. Mountjoy, C. Q., Hall, R. Lancet, 1974, ii, 415. 24. Hutton, W. N. ibid. p. 53. 25. Prange, A. J., Breese, G. R., Cott, J. M., Martin, B. R., Cooper, B. R., Wilson, I. C., Plotnikoff, N. P. Life Sci. 1974, 14, 447. 26. Breese, G. R., Cott, J. M., Cooper, B. R., Prange, A. J., Lipton, M. A. ibid. p. 1053. 27. Segal, D. S., Mandell, A. J. in The Thyroid Axis, Drugs and Behaviour (edited by A. J. Prange); p. 129. New York, 1974. 28. Brown, M., Vale, W. Endocrinology, 1975, 96, 1333. 29. Moss, R. L., McCann, S. M. Science, 1973, 181, 177. 30. Mortimer, C. H., McNeilly, A. S., Fisher, R. A., Murray, M. A. F., Besser, G. M. Br. med. J. 1974, iv, 617. 31. Dyer, R. G., Dyball, R. E. J. Nature, 1974, 252, 486. 32. Renaud, L. P., Martin, J. B. Brain Res. 1975, 86, 150. 33. Renaud, L. P., Martin, J. B., Brazeau, P. Nature, 1975, 255, 233. 34. Renaud, L. P., Martin, J. B. Brain Res. 1975, 93, 145. 35. Dyer, R. G. in Hypothalamic Hormones: Chemistry, Physiology, Pharmacology and Clinical Uses (edited by M. Motta and L. Martin). Amsterdam (in the press). 36. Cowan, W. M., Raisman, G., Powell, T. P. S. J. Neurol. Neurosurg. Psychiat. 1965, 28, 137. 37. Kievit, J., Kuypers, H. G. J. M. Science, 1974, 187, 660. 38. Szentagothai, J., Flerko, B., Mess, B., Halasz, B. Hypothalamic Control of the Anterior Pituitary; p. 22. Budapest, 1968. 39. Jackson, I. M. D., Reichlin, S. Program of the 57th Annual Meeting of Endocrine Society, June, 1975, p. 96. 40. White, W. F., Hedlund, M. T., Weber, G. F., Rippel, R. H., Johnson, E. S., Wilber, J. F. Endocrinology, 1974, 94, 1422. 41. Nir, I., Behroozi, K., Assael, M., Ivriani, I., Sulman, F. G.
Neuroendocrinology, 1969, 4, 122. 42. Reiter, R. J., Morgan, W. W., Talbot, J. A. Archs int. Pharmacodyn. 1973, 202, 219. 43. Iverson, L. L. Science, 1975, 188, 1084. 44. Labrie, F. in Neuroendocrine Relationships (edited by L. Fisher). New York (in the press). 45. Martin, J. B., Renaud, L. P., Brazeau, P. Science, 1974, 186, 538.
previous report 240 in which the P.C.A. extracts of colonic
Hypothesis
carcinomas could be separated into C.E.A.-active and i-active fractions. And it is likely that the incomplete
blood-group-like activity which was previously reported in a C.E.A. preparation 12 was caused by the presence of contaminant precursor substances. Earlier studies of the carbohydrate and aminoacid compositions of the fractions a to e of the tumour 1 C.E.A. preparation had suggested that fraction a was contaminated with a mucin-type glycoprotein.18 It was not surprising, therefore, that this fraction had the
highest blood-group precursor-like activity. The biological role of the blood-group antigens is not yet understood; nor is it known whether their deficient synthesis in entodermal tumours and in their metastases is a manifestation of rapid tissue growth, or However, the present studies indicate that the precursorlike activities of these tumours deserve detailed investigation, as does the possible release of these antigens into the bloodstream. Sensitive radioimmunoassays are currently being designed for the accurate measurement of these antigens in tumours and their fractions. Thus, high titre anti-I and anti-i cold. agglutinins which recognise several types of determinants on blood-group precursors may well prove to be powerful reagents in the detection and monitoring of a "new" set of tumour-associated antigens.
in
some
way related to tumour invasiveness.
We thank Dr M. A. Bukhari for the monosaccharide analyses; Mr S. Patel for the radioimmunoassays; and Mr R. Childs and Miss J. W. Summers for technical assistance. Requests for reprints should be addressed to T. F. REFERENCES
1. Hakomori, S., Kijimoto, S., Siddiqui, B. in Membrane Research (edited by C. F. Fox); p. 253. New York, 1972. 2. Masamune, H., Kawasaki, H., Abe, S., Oyama, K., Yamaguchi, Y. Tohoku J. exp. Med. 1958, 68, 81. 3. Iseki, S., Furukawa, K., Ishikara, K. Proc. Jap. Acad. 1962, 38, 556. 4. Hakomori, S., Koscielak, J., Bloch, H., Jeanloz, R. J. Immun. 1967, 98, 31. 5. Davidsohn, I., Ni, L. Y., Stejskal, R. Archs Path. 1971, 92, 456. 6. Gold, R., Freedman, S. O. J. exp. Med. 1965, 121, 439. 7. Gold, R., Freedman, S. O. ibid. 1965, 122, 467. 8. Lo Gerfo, P., Krupey, J., Hansen, H. J. New Engl. J. Med. 1971, 285, 138. 9. Zamchek, N., Moore, T. L., Dhar, P., Kupchik, H. Z. ibid. 1972, 286, 83. 10. Martin, F., Martin, M. S. Int. J. Cancer, 1970, 6, 352. 11. Mach, J. P., Pusztaszeri, G. Immunochemistry, 1972, 9, 1031. 12. Simmons, D. A. R., Perlmann, P. Cancer Res. 1973, 33, 313. 13. Feizi, T., Kabat, E. A., Vicari, G., Anderson, B., Marsh, W. L. J. exp. Med. 1971, 133, 39. 14. Feizi, T., Kabat, E. A., Vicari, G., Anderson, B., Marsh, W. L. J. Immun. 1971, 106, 1578. 15. Feizi, T., Kabat, E. A. J. exp. Med. 1972, 135, 1247. 16. Krupey, J., Gold, P., Freedman, S. O. ibid. 1968, 128, 387. 17. Turberville, C., Darcy, D. A., Laurence, D. J. R., Johns, E. W., Neville, A. M. Immunochemistry, 1973, 10, 841. 18. Turberville, C. Unpublished. 19. Clamp, J. R., Bhatti, T., Chambers, R. E. Meth. biochem. Analysis, 1971, 19, 229. 20. Feizi, T., Cederqvist, L. L., Childs, R. Br. J. Hœmat. 1975, 30, 485. 21. Vicari, G., Kabat, E. A. J. Immun. 1969, 102, 821. 22. Vicari, G., Kabat, E. A. Biochemistry, 1970, 9, 3414. 23. Laurence, D. J. R., Stevens, U., Bettelheim, R., Darcy, D., Leese, C., Turberville, C., Alexander, P., Johns, E. W., Neville, A. M. Br. med. J. 1972, iii, 605. 24. Cooper, A. G., Brown, M. C., Kirsh, M. E., Rule, A. H. J. Immun.
1974, 113, 1246.
Addendum T antigen (a presumed precursor of blood-group MN antigens) has recently been reported in human breast cancers (Springer, G. F., Desai, P. R., Banatwala, I. y. natn. Cancer Inst. 1975, S4, 335).
HYPOTHALAMIC PEPTIDES: NEW EVIDENCE FOR " PEPTIDERGIC " PATHWAYS IN THE C.N.S.
JOSEPH B.
MARTIN
LEO P. RENAUD
PAUL BRAZEAU
Division of Neurology, Department of Montreal General Hospital and McGill Montreal, Canada
Sum ary
Medicine, University,
Hypothalamic peptides, discovered be-
cause of their effects on stimulation or inhibition of anterior-pituitary-hormone release, have now been shown to be distributed in many regions of the central nervous system (C.N.S.). Demonstration of their localisation in nerve terminals, of behavioural effects exclusive of anterior pituitary actions, and of depressant effects in electrophysiological experiments suggests a role in central-nervous-system function. It is postulated that a system of peptidergic neurons exists within the C.N.S. with axon terminations both on the pituitary portal system and on other neurons. THE neural control of endocrine homoeostasis has been extensively investigated. The isolation and structural characterisation of several hypothalamic peptides that are known to regulate anterior-pituitary-hormone secretion has led to the emergence of evidence that points to new concepts of brain-hormone interaction, and permits speculation with respect to the role of " neurosecretion " in central-nervous-system (c.N.s.) function. We review here some of the information on the role that peptides, particularly hypothalamic peptides, may have in the C.N.s. and speculate on the existence of a system of peptide neurosecretory hypothalamic neurons that may terminate, not only on the pituitary portal capillary plexus, but also in extrahypothalamic regions of the brain. Several independent observations have drawn attention to the possibility that low-molecular-weight shortchain polypeptides have important direct effects on brain function. For example, angiotensin n, which promotes aldosterone secretion from the adrenal gland, may also be synthesised within the C.N.S.l Hypothalamic injection of angiotensin 11 causes animals in a normal state of water balance to drink, and its direct application to certain hypothalamic neurons increases their spike-discharge activity.2 Adrenocorticotrophin (A.C.T.H.) and smaller peptide fragments of A.C.T.H. that are devoid of corticotropic activity can influence the behaviour of laboratory animals 3 The c.N.s. actions of A.C.T.H. may explain certain of the behavioural alterations that accompany excessive A.C.T.H. secretion in Cushing’s disease. Low-molecularweight peptides have also been implicated in the regulation of sleepand memory.5 It is now possible to extend such observations to the recently identified hypothalamic peptides that regulate secretion of anterior-pituitary trophic hormones
(T.R.H.,
thyrotropin-releasing
hormone;
G.N.R.H.,
394
gonadotropin-releasing hormone; and somatostatin/ growth-hormone release-inhibiting hormone).6 It is believed that these peptides are synthesised by socalled releasing factor (peptidergic or tuberoinfundibular) neurosecretory neurons located in or near the medial basal hypothalamus. The peptides are thought to be secreted intermittently into the pituitary portal circulation from median-eminence response to a variety of stimuli.7
axon
terminals in
development of specific and sensitive radioimmunoassays for each of these hypothalamic peptides has permitted study of their distribution and subcellular localisation. T.R.H. is distributed widely in the medial basal hypothalamus,s whereas G.N.R.H. is localised primarily in the tuberal (arcuate) nuclei.9 Somat6statin The
is present in the arcuate and ventromedial nuclei and in the preoptic area, with lesser concentrations in other
regions of hypothalamus.
These differing patterns of distribution argue for a certain selectivity of neuronal localisation within individual hypothalamic nuclei. Immunohistochemical and biochemical studies of hypothalamic tissue have shown T.R.H.,l1 G.N.R.H.,’ and somatostatin 13,14 to be concentrated primarily within nerve terminals and synaptosomes. T.R.H. has been shown to be synthesised in vitro by hypothalamic tissue obtained from both mammalian 15 and subTissue from cerebral cortex mammalian species.16 in the rat is, however, incapable of synthesising T.R.H./5 suggesting that the localisation of synthetic mechanisms resides in the hypothalamus. The development of radioimmunoassays for the hypothalamic peptides has also resulted in the unexpected discovery that significant quantities of each peptide are present in extrahypothalamic regions, although concentrations are highest in the hypo-B thalamus. It is estimated in the rat that two-thirds of brain T.R.H. content is localised outside the hypothalamus 17; an extrahypothalamic localisation of T.R.H. has also been found in the human brain 18 G.N.R.H. is reported to be present in the cerebral cortex of the rat, and to show changes in concentration in response to stress.l9 Somatostatin is present in readily measureable quantities in the cerebral cortex, amygdala, spinal cord, and cerebrospinal fluid.2O We have shown that somatostatin within the cerebral cortex and amygdala is localised primarily within synaptosomes.21 Thus, it would appear that these peptides are located in nerve terminals in various regions of the brain. Administration of T.R.H., G.N.R.H., and somatostatin to laboratory animals and man results in behavioural effects not attributable directly to an action on the pituitary. T.R.H. has been suggested to have antidepressant properties in man,22 although there is controversy over its therapeutic efficacy.23.24 Probably related to a C.N.S. stimulatory role is the ability of T.R.H. to counteract the depressant effects of barbiturates, to antagonise ethanol narcosis, and to increase spontaneous motor activity in laboratory animals 2’ T.R.H. also reduces the potentially lethal effects of strychnine.28 G.N.R.H. restores copulatory behaviour in hypophyst.tomised rats,29 and appears to have aphrodisiac effects when administered systemically in man.3o Animal studies with somatostatin indicate that it has sedative effects and prolongs the activity of pentobarbital2s; it is also reported to potentiate the effects
of strychnine.28 Many of these effects remain after ablation (hypophysectomy, thyroidtarget-organ ectomy, or gonadectomy), indicating a significant direct That T.R.H. is present in the brain of C.N.S. action. primitive vertebrates that lack a pituitary or a thyroid 17 speaks eloquently for the primitive ancestry of the peptide and to the probability of a significant direct role in the C.N.S. Electrophysiological studies have provided evidence that polypeptides can influence neuronal responses. Microiontophoretic application of T.R.H., G.N.R.H., and somatostatin to single neurons is associated with a prominent but readily reversible decrease in excitability."-31 This effect is observed both in the hypothalamus and in several extrahypothalamic areas, including the cerebral cortex, cerebellurn, and spinal cord. The observation that only a certain population of neurons display peptide-sensitivity argues for a degree of selectivity and specificity of effect with respect to the individual peptides. If hypothalamic peptides are synthesised only by hypothalamic tissue, how do they become distributed Recent electrophysioto extrahypothalamic sites? logical studies have demonstrated evidence for axon collaterals of hypothalamic tuberoinfundibular neurons that terminate locally within the hypothalamus and also extend to brain sites quite distant from the hypothalamus-e.g., thalamic nuclei 34 Evidence of axon bifurcations of hypothalamic tuberoinfundibular neurons has been documented in anatomical studies.35 Significant hypothalamic projections to the amygdala,31 cerebral cortex,37 and brainstem 38 are known to exist. If diffusely distributed hypothalamic fibre systems can be shown to originate from neurons that produce the peptides (i.e., neurosecretory peptidergic neurons), this could explain the specific extrahypothalamic localisations of these substances. Reichlin and associates39 have reported that hypothalamic lesions in the thyroid regulatory area result in a reduction in T.R.H. content in the cerebral cortex. Taken together, these findings support the possi-
bility of a primary hypothalamic site for formation and distribution of these peptides. Activity in the tuberoinfundibular neurosecretory system could result in simultaneous release of peptides, not only into the capillary circulation in the median eminence, but also in other regions of the central nervous system that have synaptic connections from the hypothalamus. It is conceivable that release of these peptides from nerve terminals in extrahypothalamic regions of brain may subserve important biological functions such as feedback control and/or regulation of behaviour through a pre-synaptic or post-synaptic action affecting the electrical activity of distant C.N.S. neurons. In agreement with such an hypothesis for peptide modulation of neuronal excitability are the observations that all three hypothalamic peptides are now reported to be present in the pineal gland,2o,40 and that pinealectomy induces seizures in normal 41 and parathyroidectomised rats.42 Recent studies have implicated cyclic A.M.P. in central neuronal synaptic events 43 Since hypothalamic peptides act at a pituitary level by stimulating (T.R.H., G.N.R.H.) or inhibiting (somatostatin) cyclic A.M.P. formation,44. it is conceivable that this mechanism may
395 also contribute to their electrophysiological and behavioural effects in the C.N.S. The involvement of hypothalamic peptides in regulation of anterior-pituitary-hormone secretion may be a rather recent evolutionary development. This conclusion is supported by the relative lack of specificity of effect of the identified hypothalamic hormones on anterior pituitary. T.R.H. stimulates both T.S.H. and prolactin release 6; G.N.R.H. stimulates L.H. and F.S.H.44 Somatostatin in addition to inhibiting G.H. secretion also blocks T.R.H.-stimulated T.S.H. secretion and decreases insulin, glucagon, and gastrin release s The intermittent or episodic release of pituitary hormones, which is now recognised as the predominant form of hormone release." may be a reflection of underlying ongoing activity of basic C.N.S. rhythms. Homoeostasis in the periphery is accomplished by the prolonged metabolic effects of peripheral target hormones. Thus, stable hormonal control is achieved despite intermittent or rhythmic oscillations in hypothalamic neural function. We hypothesise, therefore, that hypothalamic peptidergic neurons, like monoaminergic neurons (dopaminergic, catecholaminergic, and serotoninergic) may be involved in the formation of a diffuse neural network that terminates in widespread regions of the C.N.S. The release of these peptides into the pituitary circulation may be but one facet of their biological significance. Their role in the C.N.S. may ultimately prove to be of equal or even greater importance in terms of neurobiological regulation. Proof of such a hypothesis will require further studies of the subcellular localisation of these peptides in extrahypothalamic tissue, demonstration of their release from non-hypothalamic nerve-endings after hypothalamic stimulation, and full characterisation of their putative role as either neurotransmitters or synaptic modulators. The intense interest in this question and the rapid emergence of data from several disciplines indicate that some answers will be forthcoming in the near future. Requests for reprints should be addressed to J. B. M., Division of Neurology, Montreal General Hospital, Suite 753, Livingston Hall, 1650 Cedar Avenue, Montreal, Quebe< H3G 1A4, Canada. REFERENCES 1. Reid, I. A., Ramsay, D. J., Malayan, S. A., Gänong, W. F. Program of the 57th Annual Meeting of Endocrine Society, June, 1975,
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A.
Methods and Devices AN EXTRACTOR FOR SCALPEL BLADES
J. McKIE
A. SHAW
West of Scotland Health Boards, Department of Clinical Physics and Bio-Engineering,
Glasgow ALTHOUGH it is easy to attach disposable scalpel blades handles made to British or American standardstheir removal presents problems. Some of the smaller blades to
(e.g., Swann-Morton 10A, 11, 15) are especially difficult, and the task is not made easier when the hands are gloved and wet. There is a strong likelihood of a slipped tool or a snapped blade and a cut finger. At the Royal Infirmary, Glasgow, the injury most frequently reported was back injury sustained whilst lifting patients. Second place was shared by cuts received whilst discarding scalpel blades and scratches from used hypodermic needles when the proper disposal procedure had not been followed. We have devised a tool for extracting scalpel blades from their holders. The tool (fig. 1) has two jaws moved by " scissor-lever handles. The jaws are recessed to form a channel into which the scalpel handle extension fits, and the mating
14. Pelletier,
G., Labrie, F., Arimura, A., Schally, A. V. Am. Anat. J.
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Neuroendocrinology, 1969, 4, 122. 42. Reiter, R. J., Morgan, W. W., Talbot, J. A. Archs int. Pharmacodyn. 1973, 202, 219. 43. Iverson, L. L. Science, 1975, 188, 1084. 44. Labrie, F. in Neuroendocrine Relationships (edited by L. Fisher). New York (in the press). 45. Martin, J. B., Renaud, L. P., Brazeau, P. Science, 1974, 186, 538.