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Clinical and immunological significance of neopterin measurement in cerebrospinal fluid in patients with febrile convulsions
Brain & Development 21 (1999) 458±460
Original article
www.elsevier.com/locate/braindev
Clinical and immunological signi®cance of neopterin measurement in cerebrospinal ¯uid in patients with febrile convulsions Yasuhiko Kawakami a,*, Yoshitaka Fukunaga b, Kentaro Kuwabara a, Takehisa Fujita a, Osamu Fujino a, Kiyoshi Hashimoto a a
Nippon Medical School Second Hospital, Department of Pediatrics, 1-396, Kosugi, Nakahara-ward, Kawasaki, Kanagawa 211-8533, Japan b Nippon Medical School Main Hospital, Department of Pediatrics, Tokyo 113-8603, Japan Received 12 October 1998; received in revised form 17 May 1999; accepted 17 May 1999
Abstract Neopterin is synthesized mainly by monocytes/macrophages and is considered to be a marker for activation of the cellular immune system. It has been reported that cerebrospinal ¯uid (CSF) neopterin levels are signi®cantly higher in patients with bacterial meningitis than in those with aseptic meningitis or non-pleocytotic CSF. In this study levels of neopterin and interferon-gamma (IFN-g ) were measured in children with non-pleocytotic CSF. The CSF neopterin levels were signi®cantly higher in patients with typical febrile convulsions (FCs) (15:0 ^ 4:5 nmol/l) than in those with pyrexia without convulsions (6:5 ^ 2:7 nmol/l) or convulsions without pyrexia, namely, epilepsy (4:8 ^ 2:4 nmol/ l). The CSF neopterin/serum neopterin ratio (C/S ratio) was also higher in patients with typical FCs (1:54 ^ 0:83) than in those with pyrexia without convulsions (0:32 ^ 0:18) or convulsions without pyrexia (0:77 ^ 0:28). Patients with prolonged FCs tended to have higher CSF neopterin levels than those with typical FCs. There was also a tendency for CSF IFN-g levels to be higher in patients with FCs than in those with pyrexia without convulsions or convulsions without pyrexia. The results of the present study suggest that some immune activation in the central nervous system (CNS) compartment may be related to the mechanisms of FCs. q 1999 Elsevier Science B.V. All rights reserved. Keywords: Neopterin; Cerebrospinal ¯uid; Non-pleocytotic CSF; Febrile convulsions; Interferon-gamma
1. Introduction
2. Patients and methods
Neopterin is derived from guanosine triphosphate (GTP) and produced mainly by monocytes/macrophages, so it is considered to be a marker for activation of the cellular immune system [1]. Several authors have reported that cerebrospinal ¯uid (CSF) neopterin levels are useful for the analysis of pathophysical mechanisms in various central nervous system (CNS) diseases. CSF neopterin levels are elevated in diseases such as CNS infection [2], erythrophagocytic disorders [3], and AIDS [4,5]. We have shown that CSF neopterin levels are signi®cantly higher in patients with bacterial meningitis than in those with aseptic meningitis or non-pleocytotic CSF [6]. However, no reports on the role of CSF neopterin in convulsive diseases have been made previously. We measured CSF neopterin levels in patients with pyrexia and/or convulsions. CSF interferon-gamma (IFN-g ) was also measured in some patients in each group to investigate immune conditions.
Forty patients (26 males and 14 females) were studied. CSF and sera were collected from patients attending Nippon Medical School 2nd Hospital or Tama Nagayama Hospital from September 1991 to May 1998 for clinical indications, but whose CSF were not pleocytotic. Eleven patients (seven males and four females, aged between 0.75 years and 4.66 years) were ®nally diagnosed as having febrile convulsions (FCs). All of them had symmetric generalized tonic clonic convulsions. Eight patients seizures stopped within 10 min (typical FCs), however, three out of 11 suffered convulsions lasting over 30 min (prolonged FCs). Brain imaging (MRI and/or CT scan) and EEG was performed in all patients with FCs, and no abnormal ®ndings were noted. None of them suffered non-FCs (epileptic seizures) afterwards. Twentytwo patients (15 males and seven females, aged between 6 days and 4.33 years) had pyrexia but not convulsions. Seven (four males and three females, aged between 0.33 years and 4.5 years) with convulsions but without fever were diagnosed as having epilepsy (four patients with status epilepticus). Informed consent was obtained prior to each examination.
* Corresponding author. Tel.: 1 81-44-733-5181; fax: 1 81-44-7118826.
0387-7604/99/$ - see front matter q 1999 Elsevier Science B.V. All rights reserved. PII: S03 87-7604(99)0005 2-2
Y. Kawakami et al. / Brain & Development 21 (1999) 458±460
459
Students t-test, with a level of (P , 0:05) considered to be statistically signi®cant. 3. Results 3.1. Neopterin in patients with typical FCs, pyrexia without convulsions, and convulsions without pyrexia (Fig. 1) The CSF neopterin levels in patients with typical FCs were 15:0 ^ 4:5 nmol/l. In those with pyrexia without convulsions, they were 6:5 ^ 2:7 nmol/l, and in those with convulsions without pyrexia (epilepsy) 4:8 ^ 2:4 nmol/l. The CSF neopterin levels in patients with typical FCs were signi®cantly higher than in those with pyrexia without convulsions or those with convulsions without pyrexia. The CSF neopterin/serum neopterin ratio (C/S ratio) in patients with typical FCs was 1:54 ^ 0:83, and in six out of eight patients it was above 1.00. In patients with pyrexia without convulsions, it was 0:32 ^ 0:18, and each value was below 1.00. In patients with convulsions without pyrexia it was 0:77 ^ 0:28. 3.2. CSF neopterin in patients with FCs
Fig. 1. Distribution of CSF neopterin. CSF neopterin concentrations were determined by the method described in the text. The levels were signi®cantly higher in patients with typical febrile convulsions than in those with pyrexia without convulsions, or convulsions without pyrexia (epilepsy).
Samples were kept frozen at 2708C until they were measured. Neopterin was measured in CSF and sera for all patients by the high performance liquid chromatography (HPLC) method [7]. IFN-g levels were determined by radio-immuno assay (RIA) [8]. Data were reported as mean ^ standard deviation and analyzed by unpaired
CSF neopterin levels in patients with prolonged FCs were 131, 52, 25 nmol/l (Table 1), and C/S ratio in patients with prolonged FCs was 8.2, 4.0, 3.1, respectively. CSF neopterin and C/S ratio in patients with prolonged FCs tended to be at higher levels than in those with typical FCs. 3.3. Relationship between CSF neopterin and IFN-g in patients with non-pleocytotic CSF (Table 1) CSF IFN-g was measured in 13 patients (®ve patients with FCs, ®ve with pyrexia without convulsions, and three with convulsions without pyrexia). CSF IFN-g in patients with FCs tended to be at higher levels than in those with pyrexia without convulsions or convulsions without
Table 1 Pro®les of the patients in whom both neopterin and IFN-g in CSF were measured No.
Age (years)
Clinical manifestations
Neopterin (nmol/l)
IFN-g (IU/ml)
1 2 3 4 5 6 7 8 9 10 11 12 13
0.75 1.5 1 2 3.5 0.75 0.5 1.25 0.5 1 0.5 4.5 3.75
FC a, typical (5) b FC, typical (10) FC, prolonged (60) FC, prolonged (30) FC, prolonged (30) Exanthem subitum Exanthem subitum Fever of unknown origin Fever of unknown origin Fever of unknown origin Epilepsy Epilepsy, Status epilepticus Epilepsy, Status epilepticus
18 18 131 52 25 7 4 10 7 7 6 3 2
2.3 2.4 0.1 2.8 1.8 0.8 ,0.1 0.2 0.7 0.6 ,0.1 ,0.1 ,0.1
a b
FC, febrile convulsions. Minutes are given in parentheses.
460
Y. Kawakami et al. / Brain & Development 21 (1999) 458±460
pyrexia. But there was one exception whose level of CSF IFN-g was not high in spite of the most severe clinical symptoms in FCs with an extremely high level of CSF neopterin (patient No. 3 in Table 1). 4. Discussion There have been numerous reports about CSF neopterin in correlation with various diseases. However, no authors have discussed the relationship between CSF neopterin levels and convulsive diseases. This study shows that both CSF neopterin levels and the C/S ratio are higher in patients with FCs than in those with only pyrexia or in those with only convulsions. It is known that pleocytotic CSF has a high level of neopterin [2,6]. However, it is interesting that even in patients with FCs, whose CSF are not pleocytotic, their CSF have higher levels of neopterin than those with pyrexia without convulsions or those with convulsions without pyrexia. It has been reported that relatively higher values of neopterin in CSF, as compared with serum, indicate that the main focus of impairment is the CNS [9]. Also we have shown that C/S ratios in most patients with FCs were more than 1.00. On the other hand all patients with pyrexia without convulsions were below 1.00. The CSF albumin/serum albumin ratio [10] in two patients with prolonged FCs and in three with typical FCs (data not shown) indicated their blood brain barriers (BBB) were intact. These ®ndings suggest that an immune response in the CNS compartment with patients with FCs is activated. Neopterin is known to be produced by macrophage/ monocyte [1] or endotherial cells [11] under stimulation by IFN-g . Identi®cation of the origin of macrophages and microglial cells [12,13] and the possibility of neopterin production by microglial cells [14] has been reported. We have demonstrated that CSF neopterin levels were elevated in patients with FCs, whose CSF had no pleocytosis. However, what produces neopterin in the CNS remains unknown. CSF IFN-g levels in patients with FCs did not always seem to depend on the severity or duration of the convulsions (Table 1). This suggests that IFN-g is not the only stimulant of neopterin production, as previously reported [15,16]. Even though patients had severe convulsions such as, status epilepticus, CSF neopterin or IFN-g in those with non-FCs did not increase. So we assume that some type/s of immune activation in the CNS resulting in the elevation of CSF neopterin may be associated with a pathophysiology of FCs, but not non-FCs. In animal models, it was reported that the regional GABA concentration in subcortical structures was related to the onset of FCs [17]. The present study suggests that some immune activation in the CNS may be related to the genesis of FCs. Further detailed research is needed to resolve the pathogenesis of FCs.
Acknowledgements We are grateful to Prof. Timothy D. Minton, Nippon Medical School, for reviewing the manuscript. This study was supported by a grant from the Ministry of Education of Japan. References [1] Wachter H, Fuchs D, Hausen A, Reibnegger G, Werner ER. Neopterin as marker for activation of cellular immunity: immunologic basis and clinical application. Adv Clin Chem 1989;27:81±141. [2] Fredrikson S, Eneroth P, Link H. Intrathecal production of neopterin in aseptic meningo-encephalitis and multiple sclerosis. Clin Exp Immunol 1987;67:76±81. [3] Howells DW, Strobel S, Smith I, Levinsky RJ, Hyland K. Central nervous system involvement in the erythrophagocytic disorders of infancy: the role of cerebrospinal ¯uid neopterins in their differential diagnosis and clinical management. Pediatr Res 1990;28:116±119. [4] Fuchs D, Chiodi F, Albert J, AsjoÈ B, Hagberg L, Hausen A, et al. Neopterin concentrations in cerebrospinal ¯uid and serum of individuals infected with HIV-1. AIDS 1989;3:285±288. [5] Brew BJ, Bhalla RB, Paul M, Gallardo H, McArthur JC, Schwartz MK, et al. Cerebrospinal ¯uid neopterin in human immunode®ciency virus type 1 infection. Ann Neurol 1990;28:556±560. [6] Kawakami Y, Fukunaga Y, Hashimoto K. Changes of neopterin in cerebrospinal ¯uid and serum in children with meningitis. No To Hattatsu (Tokyo) [in Japanese] 1996;28:23±29. [7] Hausen A, Fuchs D, KoÈnig K, Wachter H. Determination of neopterine in human urine by reversed phase high-performance liquid chromatography. J Chromatogr 1982;227:61±70. [8] Woloszczuk W. A sensitive immunoradiometric assay for gamma interferon, suitable for its measurement in serum [Letter]. Clin Chem 1985;31:1090. [9] Ali A, Rudge P, Dalgleish AG. Neopterin concentrations in serum and cerebrospinal ¯uid in HTLV-I infected individuals. J Neurol 1992;239:270±272. È hman S. Principles of albumin and IgG analyses [10] Tibbling G, Link H, O in neurological disorders. I. Establishment of reference values. Scand J Clin Lab Invest 1977;37:385±390. [11] Andert SE, Griesmacher A, Zuckermann A, MuÈller MM. Neopterin release from human endothelial cells is triggered by interferongamma. Clin Exp Immunol 1992;88:555±558. [12] Watkins BA, Dorn HH, Kelly WB, Armstrong RC, Potts BJ, Michaels F, et al. Speci®c tropism of HIV-1 for microglial cells in primary human brain cultures. Science 1990;249:549±553. [13] Giulian D, Vaca K, Noonan CA. Secretion of neurotoxins by mononuclear phagocytes infected with HIV-1. Science 1990;250:1593± 1596. [14] Tyor WR, Glass JD, Grif®n JW, Becker PS, McArthur JC, Bezman L, et al. Cytokine expression in the brain during the acquired immunode®ciency syndrome. Ann Neurol 1992;31:349±360. [15] Grif®n DE, Ward BJ, Jauregui E, Johnson RT, Vaisberg A. Immune activation during measles: interferon-g and neopterin in plasma and cerebrospinal ¯uid in complicated and uncomplicated disease. J Infect Dis 1990;161:449±453. [16] Grif®n DE, McArthur JC, Cornblath DR. Neopterin and interferongamma in serum and cerebrospinal ¯uid of patients with HIV-associated neurologic disease. Neurology 1991;41:69±74. [17] Morimoto T, Nagao H, Sano N, Takahashi M, Matsuda H. Hyperthermia-induced seizures with a servo system: neurophysiological roles of age, temperature elevation rate and regional GABA content in the rat. Brain Dev 1990;12:279±283.
Original article
www.elsevier.com/locate/braindev
Clinical and immunological signi®cance of neopterin measurement in cerebrospinal ¯uid in patients with febrile convulsions Yasuhiko Kawakami a,*, Yoshitaka Fukunaga b, Kentaro Kuwabara a, Takehisa Fujita a, Osamu Fujino a, Kiyoshi Hashimoto a a
Nippon Medical School Second Hospital, Department of Pediatrics, 1-396, Kosugi, Nakahara-ward, Kawasaki, Kanagawa 211-8533, Japan b Nippon Medical School Main Hospital, Department of Pediatrics, Tokyo 113-8603, Japan Received 12 October 1998; received in revised form 17 May 1999; accepted 17 May 1999
Abstract Neopterin is synthesized mainly by monocytes/macrophages and is considered to be a marker for activation of the cellular immune system. It has been reported that cerebrospinal ¯uid (CSF) neopterin levels are signi®cantly higher in patients with bacterial meningitis than in those with aseptic meningitis or non-pleocytotic CSF. In this study levels of neopterin and interferon-gamma (IFN-g ) were measured in children with non-pleocytotic CSF. The CSF neopterin levels were signi®cantly higher in patients with typical febrile convulsions (FCs) (15:0 ^ 4:5 nmol/l) than in those with pyrexia without convulsions (6:5 ^ 2:7 nmol/l) or convulsions without pyrexia, namely, epilepsy (4:8 ^ 2:4 nmol/ l). The CSF neopterin/serum neopterin ratio (C/S ratio) was also higher in patients with typical FCs (1:54 ^ 0:83) than in those with pyrexia without convulsions (0:32 ^ 0:18) or convulsions without pyrexia (0:77 ^ 0:28). Patients with prolonged FCs tended to have higher CSF neopterin levels than those with typical FCs. There was also a tendency for CSF IFN-g levels to be higher in patients with FCs than in those with pyrexia without convulsions or convulsions without pyrexia. The results of the present study suggest that some immune activation in the central nervous system (CNS) compartment may be related to the mechanisms of FCs. q 1999 Elsevier Science B.V. All rights reserved. Keywords: Neopterin; Cerebrospinal ¯uid; Non-pleocytotic CSF; Febrile convulsions; Interferon-gamma
1. Introduction
2. Patients and methods
Neopterin is derived from guanosine triphosphate (GTP) and produced mainly by monocytes/macrophages, so it is considered to be a marker for activation of the cellular immune system [1]. Several authors have reported that cerebrospinal ¯uid (CSF) neopterin levels are useful for the analysis of pathophysical mechanisms in various central nervous system (CNS) diseases. CSF neopterin levels are elevated in diseases such as CNS infection [2], erythrophagocytic disorders [3], and AIDS [4,5]. We have shown that CSF neopterin levels are signi®cantly higher in patients with bacterial meningitis than in those with aseptic meningitis or non-pleocytotic CSF [6]. However, no reports on the role of CSF neopterin in convulsive diseases have been made previously. We measured CSF neopterin levels in patients with pyrexia and/or convulsions. CSF interferon-gamma (IFN-g ) was also measured in some patients in each group to investigate immune conditions.
Forty patients (26 males and 14 females) were studied. CSF and sera were collected from patients attending Nippon Medical School 2nd Hospital or Tama Nagayama Hospital from September 1991 to May 1998 for clinical indications, but whose CSF were not pleocytotic. Eleven patients (seven males and four females, aged between 0.75 years and 4.66 years) were ®nally diagnosed as having febrile convulsions (FCs). All of them had symmetric generalized tonic clonic convulsions. Eight patients seizures stopped within 10 min (typical FCs), however, three out of 11 suffered convulsions lasting over 30 min (prolonged FCs). Brain imaging (MRI and/or CT scan) and EEG was performed in all patients with FCs, and no abnormal ®ndings were noted. None of them suffered non-FCs (epileptic seizures) afterwards. Twentytwo patients (15 males and seven females, aged between 6 days and 4.33 years) had pyrexia but not convulsions. Seven (four males and three females, aged between 0.33 years and 4.5 years) with convulsions but without fever were diagnosed as having epilepsy (four patients with status epilepticus). Informed consent was obtained prior to each examination.
* Corresponding author. Tel.: 1 81-44-733-5181; fax: 1 81-44-7118826.
0387-7604/99/$ - see front matter q 1999 Elsevier Science B.V. All rights reserved. PII: S03 87-7604(99)0005 2-2
Y. Kawakami et al. / Brain & Development 21 (1999) 458±460
459
Students t-test, with a level of (P , 0:05) considered to be statistically signi®cant. 3. Results 3.1. Neopterin in patients with typical FCs, pyrexia without convulsions, and convulsions without pyrexia (Fig. 1) The CSF neopterin levels in patients with typical FCs were 15:0 ^ 4:5 nmol/l. In those with pyrexia without convulsions, they were 6:5 ^ 2:7 nmol/l, and in those with convulsions without pyrexia (epilepsy) 4:8 ^ 2:4 nmol/l. The CSF neopterin levels in patients with typical FCs were signi®cantly higher than in those with pyrexia without convulsions or those with convulsions without pyrexia. The CSF neopterin/serum neopterin ratio (C/S ratio) in patients with typical FCs was 1:54 ^ 0:83, and in six out of eight patients it was above 1.00. In patients with pyrexia without convulsions, it was 0:32 ^ 0:18, and each value was below 1.00. In patients with convulsions without pyrexia it was 0:77 ^ 0:28. 3.2. CSF neopterin in patients with FCs
Fig. 1. Distribution of CSF neopterin. CSF neopterin concentrations were determined by the method described in the text. The levels were signi®cantly higher in patients with typical febrile convulsions than in those with pyrexia without convulsions, or convulsions without pyrexia (epilepsy).
Samples were kept frozen at 2708C until they were measured. Neopterin was measured in CSF and sera for all patients by the high performance liquid chromatography (HPLC) method [7]. IFN-g levels were determined by radio-immuno assay (RIA) [8]. Data were reported as mean ^ standard deviation and analyzed by unpaired
CSF neopterin levels in patients with prolonged FCs were 131, 52, 25 nmol/l (Table 1), and C/S ratio in patients with prolonged FCs was 8.2, 4.0, 3.1, respectively. CSF neopterin and C/S ratio in patients with prolonged FCs tended to be at higher levels than in those with typical FCs. 3.3. Relationship between CSF neopterin and IFN-g in patients with non-pleocytotic CSF (Table 1) CSF IFN-g was measured in 13 patients (®ve patients with FCs, ®ve with pyrexia without convulsions, and three with convulsions without pyrexia). CSF IFN-g in patients with FCs tended to be at higher levels than in those with pyrexia without convulsions or convulsions without
Table 1 Pro®les of the patients in whom both neopterin and IFN-g in CSF were measured No.
Age (years)
Clinical manifestations
Neopterin (nmol/l)
IFN-g (IU/ml)
1 2 3 4 5 6 7 8 9 10 11 12 13
0.75 1.5 1 2 3.5 0.75 0.5 1.25 0.5 1 0.5 4.5 3.75
FC a, typical (5) b FC, typical (10) FC, prolonged (60) FC, prolonged (30) FC, prolonged (30) Exanthem subitum Exanthem subitum Fever of unknown origin Fever of unknown origin Fever of unknown origin Epilepsy Epilepsy, Status epilepticus Epilepsy, Status epilepticus
18 18 131 52 25 7 4 10 7 7 6 3 2
2.3 2.4 0.1 2.8 1.8 0.8 ,0.1 0.2 0.7 0.6 ,0.1 ,0.1 ,0.1
a b
FC, febrile convulsions. Minutes are given in parentheses.
460
Y. Kawakami et al. / Brain & Development 21 (1999) 458±460
pyrexia. But there was one exception whose level of CSF IFN-g was not high in spite of the most severe clinical symptoms in FCs with an extremely high level of CSF neopterin (patient No. 3 in Table 1). 4. Discussion There have been numerous reports about CSF neopterin in correlation with various diseases. However, no authors have discussed the relationship between CSF neopterin levels and convulsive diseases. This study shows that both CSF neopterin levels and the C/S ratio are higher in patients with FCs than in those with only pyrexia or in those with only convulsions. It is known that pleocytotic CSF has a high level of neopterin [2,6]. However, it is interesting that even in patients with FCs, whose CSF are not pleocytotic, their CSF have higher levels of neopterin than those with pyrexia without convulsions or those with convulsions without pyrexia. It has been reported that relatively higher values of neopterin in CSF, as compared with serum, indicate that the main focus of impairment is the CNS [9]. Also we have shown that C/S ratios in most patients with FCs were more than 1.00. On the other hand all patients with pyrexia without convulsions were below 1.00. The CSF albumin/serum albumin ratio [10] in two patients with prolonged FCs and in three with typical FCs (data not shown) indicated their blood brain barriers (BBB) were intact. These ®ndings suggest that an immune response in the CNS compartment with patients with FCs is activated. Neopterin is known to be produced by macrophage/ monocyte [1] or endotherial cells [11] under stimulation by IFN-g . Identi®cation of the origin of macrophages and microglial cells [12,13] and the possibility of neopterin production by microglial cells [14] has been reported. We have demonstrated that CSF neopterin levels were elevated in patients with FCs, whose CSF had no pleocytosis. However, what produces neopterin in the CNS remains unknown. CSF IFN-g levels in patients with FCs did not always seem to depend on the severity or duration of the convulsions (Table 1). This suggests that IFN-g is not the only stimulant of neopterin production, as previously reported [15,16]. Even though patients had severe convulsions such as, status epilepticus, CSF neopterin or IFN-g in those with non-FCs did not increase. So we assume that some type/s of immune activation in the CNS resulting in the elevation of CSF neopterin may be associated with a pathophysiology of FCs, but not non-FCs. In animal models, it was reported that the regional GABA concentration in subcortical structures was related to the onset of FCs [17]. The present study suggests that some immune activation in the CNS may be related to the genesis of FCs. Further detailed research is needed to resolve the pathogenesis of FCs.
Acknowledgements We are grateful to Prof. Timothy D. Minton, Nippon Medical School, for reviewing the manuscript. This study was supported by a grant from the Ministry of Education of Japan. References [1] Wachter H, Fuchs D, Hausen A, Reibnegger G, Werner ER. Neopterin as marker for activation of cellular immunity: immunologic basis and clinical application. Adv Clin Chem 1989;27:81±141. [2] Fredrikson S, Eneroth P, Link H. Intrathecal production of neopterin in aseptic meningo-encephalitis and multiple sclerosis. Clin Exp Immunol 1987;67:76±81. [3] Howells DW, Strobel S, Smith I, Levinsky RJ, Hyland K. Central nervous system involvement in the erythrophagocytic disorders of infancy: the role of cerebrospinal ¯uid neopterins in their differential diagnosis and clinical management. Pediatr Res 1990;28:116±119. [4] Fuchs D, Chiodi F, Albert J, AsjoÈ B, Hagberg L, Hausen A, et al. Neopterin concentrations in cerebrospinal ¯uid and serum of individuals infected with HIV-1. AIDS 1989;3:285±288. [5] Brew BJ, Bhalla RB, Paul M, Gallardo H, McArthur JC, Schwartz MK, et al. Cerebrospinal ¯uid neopterin in human immunode®ciency virus type 1 infection. Ann Neurol 1990;28:556±560. [6] Kawakami Y, Fukunaga Y, Hashimoto K. Changes of neopterin in cerebrospinal ¯uid and serum in children with meningitis. No To Hattatsu (Tokyo) [in Japanese] 1996;28:23±29. [7] Hausen A, Fuchs D, KoÈnig K, Wachter H. Determination of neopterine in human urine by reversed phase high-performance liquid chromatography. J Chromatogr 1982;227:61±70. [8] Woloszczuk W. A sensitive immunoradiometric assay for gamma interferon, suitable for its measurement in serum [Letter]. Clin Chem 1985;31:1090. [9] Ali A, Rudge P, Dalgleish AG. Neopterin concentrations in serum and cerebrospinal ¯uid in HTLV-I infected individuals. J Neurol 1992;239:270±272. È hman S. Principles of albumin and IgG analyses [10] Tibbling G, Link H, O in neurological disorders. I. Establishment of reference values. Scand J Clin Lab Invest 1977;37:385±390. [11] Andert SE, Griesmacher A, Zuckermann A, MuÈller MM. Neopterin release from human endothelial cells is triggered by interferongamma. Clin Exp Immunol 1992;88:555±558. [12] Watkins BA, Dorn HH, Kelly WB, Armstrong RC, Potts BJ, Michaels F, et al. Speci®c tropism of HIV-1 for microglial cells in primary human brain cultures. Science 1990;249:549±553. [13] Giulian D, Vaca K, Noonan CA. Secretion of neurotoxins by mononuclear phagocytes infected with HIV-1. Science 1990;250:1593± 1596. [14] Tyor WR, Glass JD, Grif®n JW, Becker PS, McArthur JC, Bezman L, et al. Cytokine expression in the brain during the acquired immunode®ciency syndrome. Ann Neurol 1992;31:349±360. [15] Grif®n DE, Ward BJ, Jauregui E, Johnson RT, Vaisberg A. Immune activation during measles: interferon-g and neopterin in plasma and cerebrospinal ¯uid in complicated and uncomplicated disease. J Infect Dis 1990;161:449±453. [16] Grif®n DE, McArthur JC, Cornblath DR. Neopterin and interferongamma in serum and cerebrospinal ¯uid of patients with HIV-associated neurologic disease. Neurology 1991;41:69±74. [17] Morimoto T, Nagao H, Sano N, Takahashi M, Matsuda H. Hyperthermia-induced seizures with a servo system: neurophysiological roles of age, temperature elevation rate and regional GABA content in the rat. Brain Dev 1990;12:279±283.