Plasma Tryptophan in Migraine S. Salmon, M. Fanciullacci, M. Bonciani, and F. Sicuteri Department of Clinical Pharmacology, Headache Centre, University of Florence, Italy. (Drs. Salmon, Fanciullacci, Bonciani, Sicuteri) Preliminary Report presented at the 18th Annual Headache Meeting of the American Association for the Study of Headache, Dallas, June, 1976. Reprint requests to: University of Florence, Dept. of Clinical Pharmacology, Clinica Medica Generale, 85, Viale Morgagni, 50134 Florence Italy (Dr. Salmon) Accepted for publication: 11/3/77 SYNOPSIS Free and total plasma tryptophan (TRY) was evaluated in migraine and daily headache sufferers. In migraineurs, free plasma TRY was significantly higher on the day of the attack, the day preceding and subsequent to the attack in comparison with controls. The possibility that in migraineurs a lowering of brain 5-HT may displace TRY from its binding to plasma albumin through an unknown feedback mechanism is suggested. In daily headache sufferers total plasma TRY levels were statistically significantly lower compared with controls. This may reflect a deficiency of 5-HT in the brain. (Headache 17: 238-241, 1978) "NON-ORGANIC central pain" (NOCP) includes painful conditions which are related to a functional disorder of the central nociceptive system.1,2 The most common and important expression of NOCP is idiopathic headache (IH).3 Serotonin (5-HT) is considered to act centrally by inhibiting the nociceptive system;4-8 a deficiency of 5-HT in the antinociceptive system is proposed to account for IH.9,10,1 5-HT precursors improve IH by correcting 5-HT deficiency.11 Ergotamine and the 5-HT partial agonist, methysergide, and ORG 94, can improve IH by facilitating the antinociceptive action of 5-HT.12 Plasma free tryptophan (TRY), the fraction not bound to albumin, apparently controls brain 5-HT turnover.13-15 Therefore, modifiers of the equilibrium between free and bound TRY in plasma may also influence brain 5-HT turnover.16 We studied free and total TRY plasma levels in migraine patients during attacks and in daily headache sufferers. We also report the TRY plasma variations induced by ergotamine, fenfluramine and nitroglycerin which are able to interrupt or provoke migraine. Fenfluramine, an amphetamine derivative, precipitates headache probably by its action on brain 5-HT.17-19 PATIENTS AND METHODS Fifty-one patients were studied. The migraine group consisted of 11 females and 4 males ranging from 27 to 57 years (mean ± SE = 41 ± 2.4) who suffered from severe and frequent attacks. Plasma TRY levels were estimated during attacks (30 measurements) and in headache free periods (42 measurements). Migraine attacks included the day or days of the pain, the day preceding and subsequent to the attack. Plasma TRY levels were measured in daily headache sufferers (15 females and 5 males) ranging in age from 23 to 61 years (mean ± SE = 41 ± 2.2). Ergotamine, analgesics or other drugs were withheld for one week before and during the period of this investigation. The controls were healthy, drug-free volunteers (6 females and 9 males) ranging in age from 25 to 59 years (mean ± SE = 35.8 ± 2.4). Blood samples (30 ml) were drawn early in the morning from a forearm vein using plastic syringes. All subjects had been fasting overnight. Pharmacologic investigations were carried out in another group of daily headache sufferers (9 females and 7 males). Drugs were administered on different days at 8:00 a.m., and patients were kept resting during the period of study. Ergotamine tartrate (0.250 mg) or fenfluramine hydrochloride (30 mg) was diluted in 10 ml of saline and slowly injected intravenously (2 minutes) in two groups of 5 patients. Plasma TRY levels were measured before and after drug administration at: 1) Ergotamine: 0, 30, 60, 120, 180, 240 minutes and 24 hours. 2) Fenfluramine: 0, 10, 20, 30, 60, 120 minutes and 24 hours.
3) Nitroglycerin: 0, 30, 90, 150, 210 minutes and 24 hours. The method used was that of Eccleston, with an ultrafiltration technique for the separation of free TRY and a modification of Denckla and Dewey's original method.20 The Mann Whitney U test was used for statistical comparisons when the variances of the populations were not homogeneous. RESULTS Mean free plasma TRY concentration is significantly higher during migraine when compared to mean values of controls (Z = 2.18; a < 0.02). Mean value of free as percent of total TRY is also increased (t = 5.66; p < 0.001) (Fig. 1, Table 1). Mean total plasma TRY is significantly lower in daily headache sufferers in comparison to normal subjects (U = 71.5; a < 0.02). Mean value of free as percent of total TRY is significantly increased (t = 2.19; p < 0.05) (Fig. 2, Table 1). Modifications by the three different drugs did not produce significant differences between mean values of single groups when evaluated by student t test. We emphasize that in three of six subjects, 90 minutes after administration of nitroglycerin with subsequent attacks, free plasma TRY increased and total plasma TRY decreased immediately after cessation of the induced headache (Fig. 3); in other patients, where the drugs did not provoke an attack, no changes of TRY were observed. DISCUSSION The increase of free plasma TRY in migraine previously noted in preliminary reports,21,22 is statistically significant.
This increase could be due to fasting because migraineurs usually do not eat or change their dietary habits during attacks. Diet influences brain TRY metabolism; fasted rats have higher free plasma TRY levels than rats normally fed.23 In humans, diet also seems to influence brain TRY metabolism.24 However, some migraineurs showed an increase of free plasma TRY even on the day preceding attacks when they were not likely to be fasting or changing their habitual food. An alternative interpretation is that during migraine an acute lowering of brain 5-HT could displace TRY from its binding sites to plasma protein, through an unknown
Free TRY (µg/ml)
Table 1 Total and Free Plasma Tryptophan Controls Migraine Daily Headache (n = 15) (n = 15) (n = 20) Headache-free Attack 1.26 ± 0.12 1.73 ± 0.16 1.93 ± 0.19* 1.39 ± 2.02
Total TRY (µg/ml) 12.75 ± 1.16 Free TRY as % of total 10.50 ± 1.97 Key: All values are means ± SE * = significantly different from controls In brackets the number of subjects
11.52 ± 0.65 13.99 ± 1.97
10.13 ± 0.56 20.49 ± 1.57*
8.95 ± 0.51* 16.20 ± 1.69*
feedback mechanism. This might result in making more tryptophan available to the brain, thereby correcting the 5-HT deficiency. The lowered total plasma TRY level in daily headache patients may be an adaptative mechanism to a reduced central 5-HT turnover. In support of this hypothesis could be the reduction of monoamine oxidase activity in platelets25 and free plasma 5-HT26,27 observed in daily headache sufferers. It may also be that the lowered plasma total TRY reflects a genetic deficiency of 5-HT in the brain of chronic headache sufferers. The results of this study can, therefore, be considered supportive of the central origin of daily headache as a disease of the brain 5-HT antinociceptive system, and might justify headache therapy with 5-HT precursor.28,11 Drugs which influence headache, such as fenfluramine, nitroglycerin and ergotamine can modify free and total tryptophan occasionally, but no statistically significant changes have been found on examination of the subjects. ACKNOWLEDGEMENTS Thanks to Mr. Vittorio Vivoli for expert technical assistance and to Miss Annie C. Lang for help in the translation of the manuscript. Supported by a grant from the National Research Council, Rome, Italy. REFERENCES 1.
Sicuteri F, Fanciullacci M, Anselmi B: The serotonin (5-HT) theory of migraine. Advances in Neurology 4:383-393, Raven Press, 1974.
2.
Sicuteri F: Migraine a central biochemical dysnociception. Headache 16:145-159, 1976.
3.
Sicuteri F: Pain syndrome in man following treatment with p-chlorophenylalanine. Pharm Res Comm 3:401-407, 1971.
4.
Harvey JA, Lints CE: Lesion in the medial forebrain bundle: delayed effects on sensitivity to electric shock. Science 148:250-252, 1965.
5.
Harvey JA, Schlosberg AJ, Yunger LM: Behavioural correlates of serotonin depletion. Fed Proc 34:1796-1801, 1975.
6.
Lytle LD, Messing RB, Fisher L, Pmebus L: Effects of long term corn consumption on brain serotonin and the response to electric shock. Science 190:692-694, 1975.
7.
Semanin R, Gumulka W, Valzelli L: Reduced effect of morphine in mid-brain raphe lesioned rats. Europ J Pharmacol 10:339-343, 1970.
8.
Tenen SS: The effects of p-chlorophenylalanine, a serotonin depletor on avoidance, acquisition, pain sensitivity and related behaviour in rats. Psychopharmacologia (Berl) 10:204-219, 1967.
9.
Sicuteri F: Headache biochemistry and pharmacology. Arch de Neurobiol (Madrid) 37:27-65, 1974.
10.
Sicuteri F, Anselmi B, Del Bianco PL: 5-hydroxytryptamine supersensitivity and a new theory of headache and central pain: clinical pharmacological approach with p-chlorophenylalanine. Psychopharmacologia (Berl) 20:347-356, 1973.
11.
Sicuteri F: The ingestion of serotonin precursors (l-5 hydroxytroptophan and l-typtophan) improves migraine headache. Headache 13:19-22, 1973.
12.
Fanciullacci M, Franchi G, Sicuteri F: Ergotamine and methysergide as serotonin partial agonists in migraine. Headache 16:226-231, 1976.
13.
Fernstrom JD, Wurtman RJ: Brain serotonin content: physiological dependence on plasma tryptophan levels. Science 173:149-151, 1971.
14.
Tagliamonte A, Biggio G, Geesa GL: Possible role of "free" plasma tryptophan in controlling brain tryptophan concentration. Farmacol Ter 11:251-255, 1971.
15.
Tagliamonte A, Biggio G, Vargu L, Gessa GL: Free tryptophan in serum controls brain tryptophan level and serotonin synthesis. Life Science 12:277-287, 1973.
16.
Cruzon G: Availability of tryptophan to the brain and some hormonal and drug influences on it. Advances in Biochem Psychopharmacol 10:263-271, (Raven Press), 1974.
17.
Costa E, Groppetti A, Revualta A: Action of fenfluramine on monoamine stores of rat tissues. Brit J Pharmacol 41:57-66, 1971.
18.
Del Bene E, Sicuteri F, Anselmi B, Del Bianco PL: Cefalea e fenfluramina; implicazioni farmacocliniche. B S I B S 51:1684-1688, 1975.
19.
Tagliamonte A, Tagliamonte P, Perez-Cruet J, Stern S, Gessa GL: Effect of psychotropic drugs on tryptophan concentration in the rat brain. J Pharmacol Exp Ther 177:475-480, 1971.
20.
Eccleston EG: A method for the estimation of free and total acid soluble plasma tryptophan using ultra-filtration technique. Coin Chim Acta 48:269-272, 1973.
21.
Fanciullacci M, Salmon S, Bonciani M, Sicuteri F: Free and total plasma in idiopathic headache. In: Proc Italo-Scandinavia Headache Societies Headache '76 Congress. Florence June, 1976, (in press-Difme, Torino).
22.
Salmon S, Fanciullacci M, Bonciani M, Sicuteri F: LiveIli plasmatici di triptofano libero e totale in corso di attacco emicranico. B S I B S 12:1589-1593, 1976.
23.
Culley WJ, Saunders RN, Mertz ET, Jolly DH:L Effect of a tryptophan deficient diet on brain serotonin and plasma tryptophan level. Proc Soc Exp Med 113:645-648, 1963.
24.
Perez-Cruet J, Chase TN, Murphy DL: Dietary regulation of brain tryptophan metabolism by plasma ratio of the free tryptophan and neutral amono acids in humans. Nature 248:693-695, 1974.
25.
Sicuteri F, Buffoni F, Anselmi B, Del Bianco PL: An enzyme (MAO) defect on the platelets in migraine. Res Clin Stud Headache 3:245-251, 1972.
26.
Salmon S, Del Meglio A: Progress in evaluation of free plasma and total blood 5-hydroxytryptamine. B S I B S. (in press, 1976).
27.
Somerville BW: Platelet-bound and free serotonin levels in jugular and forearm venous blood during migraine. Neurology 26:41-45, 1976.
28.
Sicuteri F: 5-hydroxytryptophan in the prophylaxis of migraine. Pharm Res Comm 4:213-218, 1972.
3) Nitroglycerin: 0, 30, 90, 150, 210 minutes and 24 hours. The method used was that of Eccleston, with an ultrafiltration technique for the separation of free TRY and a modification of Denckla and Dewey's original method.20 The Mann Whitney U test was used for statistical comparisons when the variances of the populations were not homogeneous. RESULTS Mean free plasma TRY concentration is significantly higher during migraine when compared to mean values of controls (Z = 2.18; a < 0.02). Mean value of free as percent of total TRY is also increased (t = 5.66; p < 0.001) (Fig. 1, Table 1). Mean total plasma TRY is significantly lower in daily headache sufferers in comparison to normal subjects (U = 71.5; a < 0.02). Mean value of free as percent of total TRY is significantly increased (t = 2.19; p < 0.05) (Fig. 2, Table 1). Modifications by the three different drugs did not produce significant differences between mean values of single groups when evaluated by student t test. We emphasize that in three of six subjects, 90 minutes after administration of nitroglycerin with subsequent attacks, free plasma TRY increased and total plasma TRY decreased immediately after cessation of the induced headache (Fig. 3); in other patients, where the drugs did not provoke an attack, no changes of TRY were observed. DISCUSSION The increase of free plasma TRY in migraine previously noted in preliminary reports,21,22 is statistically significant.
This increase could be due to fasting because migraineurs usually do not eat or change their dietary habits during attacks. Diet influences brain TRY metabolism; fasted rats have higher free plasma TRY levels than rats normally fed.23 In humans, diet also seems to influence brain TRY metabolism.24 However, some migraineurs showed an increase of free plasma TRY even on the day preceding attacks when they were not likely to be fasting or changing their habitual food. An alternative interpretation is that during migraine an acute lowering of brain 5-HT could displace TRY from its binding sites to plasma protein, through an unknown
Free TRY (µg/ml)
Table 1 Total and Free Plasma Tryptophan Controls Migraine Daily Headache (n = 15) (n = 15) (n = 20) Headache-free Attack 1.26 ± 0.12 1.73 ± 0.16 1.93 ± 0.19* 1.39 ± 2.02
Total TRY (µg/ml) 12.75 ± 1.16 Free TRY as % of total 10.50 ± 1.97 Key: All values are means ± SE * = significantly different from controls In brackets the number of subjects
11.52 ± 0.65 13.99 ± 1.97
10.13 ± 0.56 20.49 ± 1.57*
8.95 ± 0.51* 16.20 ± 1.69*
feedback mechanism. This might result in making more tryptophan available to the brain, thereby correcting the 5-HT deficiency. The lowered total plasma TRY level in daily headache patients may be an adaptative mechanism to a reduced central 5-HT turnover. In support of this hypothesis could be the reduction of monoamine oxidase activity in platelets25 and free plasma 5-HT26,27 observed in daily headache sufferers. It may also be that the lowered plasma total TRY reflects a genetic deficiency of 5-HT in the brain of chronic headache sufferers. The results of this study can, therefore, be considered supportive of the central origin of daily headache as a disease of the brain 5-HT antinociceptive system, and might justify headache therapy with 5-HT precursor.28,11 Drugs which influence headache, such as fenfluramine, nitroglycerin and ergotamine can modify free and total tryptophan occasionally, but no statistically significant changes have been found on examination of the subjects. ACKNOWLEDGEMENTS Thanks to Mr. Vittorio Vivoli for expert technical assistance and to Miss Annie C. Lang for help in the translation of the manuscript. Supported by a grant from the National Research Council, Rome, Italy. REFERENCES 1.
Sicuteri F, Fanciullacci M, Anselmi B: The serotonin (5-HT) theory of migraine. Advances in Neurology 4:383-393, Raven Press, 1974.
2.
Sicuteri F: Migraine a central biochemical dysnociception. Headache 16:145-159, 1976.
3.
Sicuteri F: Pain syndrome in man following treatment with p-chlorophenylalanine. Pharm Res Comm 3:401-407, 1971.
4.
Harvey JA, Lints CE: Lesion in the medial forebrain bundle: delayed effects on sensitivity to electric shock. Science 148:250-252, 1965.
5.
Harvey JA, Schlosberg AJ, Yunger LM: Behavioural correlates of serotonin depletion. Fed Proc 34:1796-1801, 1975.
6.
Lytle LD, Messing RB, Fisher L, Pmebus L: Effects of long term corn consumption on brain serotonin and the response to electric shock. Science 190:692-694, 1975.
7.
Semanin R, Gumulka W, Valzelli L: Reduced effect of morphine in mid-brain raphe lesioned rats. Europ J Pharmacol 10:339-343, 1970.
8.
Tenen SS: The effects of p-chlorophenylalanine, a serotonin depletor on avoidance, acquisition, pain sensitivity and related behaviour in rats. Psychopharmacologia (Berl) 10:204-219, 1967.
9.
Sicuteri F: Headache biochemistry and pharmacology. Arch de Neurobiol (Madrid) 37:27-65, 1974.
10.
Sicuteri F, Anselmi B, Del Bianco PL: 5-hydroxytryptamine supersensitivity and a new theory of headache and central pain: clinical pharmacological approach with p-chlorophenylalanine. Psychopharmacologia (Berl) 20:347-356, 1973.
11.
Sicuteri F: The ingestion of serotonin precursors (l-5 hydroxytroptophan and l-typtophan) improves migraine headache. Headache 13:19-22, 1973.
12.
Fanciullacci M, Franchi G, Sicuteri F: Ergotamine and methysergide as serotonin partial agonists in migraine. Headache 16:226-231, 1976.
13.
Fernstrom JD, Wurtman RJ: Brain serotonin content: physiological dependence on plasma tryptophan levels. Science 173:149-151, 1971.
14.
Tagliamonte A, Biggio G, Geesa GL: Possible role of "free" plasma tryptophan in controlling brain tryptophan concentration. Farmacol Ter 11:251-255, 1971.
15.
Tagliamonte A, Biggio G, Vargu L, Gessa GL: Free tryptophan in serum controls brain tryptophan level and serotonin synthesis. Life Science 12:277-287, 1973.
16.
Cruzon G: Availability of tryptophan to the brain and some hormonal and drug influences on it. Advances in Biochem Psychopharmacol 10:263-271, (Raven Press), 1974.
17.
Costa E, Groppetti A, Revualta A: Action of fenfluramine on monoamine stores of rat tissues. Brit J Pharmacol 41:57-66, 1971.
18.
Del Bene E, Sicuteri F, Anselmi B, Del Bianco PL: Cefalea e fenfluramina; implicazioni farmacocliniche. B S I B S 51:1684-1688, 1975.
19.
Tagliamonte A, Tagliamonte P, Perez-Cruet J, Stern S, Gessa GL: Effect of psychotropic drugs on tryptophan concentration in the rat brain. J Pharmacol Exp Ther 177:475-480, 1971.
20.
Eccleston EG: A method for the estimation of free and total acid soluble plasma tryptophan using ultra-filtration technique. Coin Chim Acta 48:269-272, 1973.
21.
Fanciullacci M, Salmon S, Bonciani M, Sicuteri F: Free and total plasma in idiopathic headache. In: Proc Italo-Scandinavia Headache Societies Headache '76 Congress. Florence June, 1976, (in press-Difme, Torino).
22.
Salmon S, Fanciullacci M, Bonciani M, Sicuteri F: LiveIli plasmatici di triptofano libero e totale in corso di attacco emicranico. B S I B S 12:1589-1593, 1976.
23.
Culley WJ, Saunders RN, Mertz ET, Jolly DH:L Effect of a tryptophan deficient diet on brain serotonin and plasma tryptophan level. Proc Soc Exp Med 113:645-648, 1963.
24.
Perez-Cruet J, Chase TN, Murphy DL: Dietary regulation of brain tryptophan metabolism by plasma ratio of the free tryptophan and neutral amono acids in humans. Nature 248:693-695, 1974.
25.
Sicuteri F, Buffoni F, Anselmi B, Del Bianco PL: An enzyme (MAO) defect on the platelets in migraine. Res Clin Stud Headache 3:245-251, 1972.
26.
Salmon S, Del Meglio A: Progress in evaluation of free plasma and total blood 5-hydroxytryptamine. B S I B S. (in press, 1976).
27.
Somerville BW: Platelet-bound and free serotonin levels in jugular and forearm venous blood during migraine. Neurology 26:41-45, 1976.
28.
Sicuteri F: 5-hydroxytryptophan in the prophylaxis of migraine. Pharm Res Comm 4:213-218, 1972.
