- Email: [email protected]
No acute suppression of cerebrospinal fluid corticotropin-releasing hormone in man by cortisol administration
Psychiatry Research 210 (2013) 662–664
Contents lists available at ScienceDirect
Psychiatry Research journal homepage: www.elsevier.com/locate/psychres
Brief report
No acute suppression of cerebrospinal fluid corticotropin-releasing hormone in man by cortisol administration Michael Kellner a,n, Cornelie Salzwedel b, Viola Wortmann a, Tatiana Urbanowicz a, Kai Boelmans a, Alexander Yassouridis c, Günter K. Stalla c, Klaus Wiedemann a a
University Hospital Hamburg-Eppendorf, Department of Psychiatry and Psychotherapy, Hamburg, Germany University Hospital Hamburg-Eppendorf, Department of Anesthesiology, Hamburg, Germany c Max Planck Institute of Psychiatry, Munich, Germany b
art ic l e i nf o
a b s t r a c t
Article history: Received 9 January 2013 Received in revised form 18 June 2013 Accepted 7 July 2013
Corticotropin-releasing hormone (CRH) in cerebrospinal fluid (CSF) is regarded as index of brain endocrine and behavioral functioning. We investigated the acute effects of intravenous cortisol (100 mg) vs. placebo on serial CSF CRH in ten healthy men. CSF CRH concentrations were not significantly suppressed by cortisol within 3 h. The origin and regulation of CSF CRH need further research. & 2013 Elsevier Ireland Ltd. All rights reserved.
Keywords: Corticotropin-releasing hormone (CRH) Cortisol Cerebrospinal fluid (CSF)
1. Introduction Corticotropin-releasing hormone (CRH) has a major role in stress response. CRH neurons from the hypothalamic paraventricular nucleus (PVN) control hypothalamic-pituitary-adrenocortical (HPA) activity. Furthermore, extra-hypothalamic CRH has been found in the brain in cortical, limbic (central nucleus of the amygdala, bed nucleus of the stria terminalis, hippocampus) and brainstem regions, where it modulates autonomic and behavioral functions, such as anxiety, food intake, sleep, arousal, locomotion, learning and memory. Therefore, CRH and HPA system dysfunctions have been hypothesized to be implicated in the pathophysiology of psychiatric diseases, particularly of mood and anxiety disorders (deKloet et al., 2005). CRH concentrations have been measured in cerebrospinal fluid (CSF) of human subjects as a putative index of central nervous system CRH functioning (Geracioti et al., 1992). However, the source and regulation of CSF CRH have not been sufficiently clarified. Following bilateral ablation of the PVN, CSF CRH declined in rats suggesting that a considerable portion may be hypothalamic (Hong et al., 1993). In monkeys intracerebroventricular CRH stimulated the pituitary–adrenocortical axis (Rock et al., 1984).
n Correspondence to: University Hospital Hamburg-Eppendorf, Dept. of Psychiatry and Psychotherapy, Martinistrasse 52, W37, D-20246 Hamburg, Germany. Tel.: +49 40 7410 52234; fax: +49 40 7410 53461. E-mail address: [email protected] (M. Kellner).
0165-1781/$ - see front matter & 2013 Elsevier Ireland Ltd. All rights reserved. http://dx.doi.org/10.1016/j.psychres.2013.07.003
However, the diurnal rhythms of CSF CRH and of plasma pituitaryadrenocortical hormones in non-human primates were not closely linked (Garrick et al., 1987; Kalin et al., 1987); despite HPA activation by physiostigmine, metyrapone (Kalin et al., 1987) and m-chlorophenylpiperazine (Garrick et al., 1987) no consecutive changes in CSF CRH were observed. Thus, CSF CRH may rather reflect extra-hypothalamic than hypophysiotropic brain functions of this peptide. Also in man lack of synchrony between lumbar CSF CRH concentrations and pituitary–adrenocortical activity was demonstrated in normal volunteers and depressed patients (Geracioti et al., 1992, 1997). In contrast, Kling et al. (1994) reported significant negative correlations of CSF CRH and simultaneous plasma cortisol in man and suggested partial glucocorticoid suppressibility of CRH secretion into CSF. Complementarily, both Newport et al. (2003) and Lee et al. (2012) found that the CRH concentration in human CSF was inversely correlated with pituitary responsiveness to exogenous CRH. Kasckow et al. (2001) observed that after feeding CSF CRH diminished in man, which may be caused by rises in plasma cortisol and associated negative feedback effects. On the other hand, Vythilingam et al. (2000) reported that the hypothalamic CRH-releaser naloxone increased plasma cortisol, but no concomitant change of CSF CRH was seen in healthy humans. Interestingly, so far the acute effects of cortisol administration on CSF CRH have not been investigated. Therefore, we performed a respective pilot study in healthy human volunteers using a serial CSF sampling technique and hypothesized suppressive effects of cortisol on CSF CRH.
M. Kellner et al. / Psychiatry Research 210 (2013) 662–664
Placebo
2. Methods
cortisol
140
CSF-Cortisol (ng/ml)
2.1. Participants Ten young men (mean age 22.7 years, range 21–27; mean body mass index 24.3 kg/m2, range 22.3–27.5) were studied. They all were non-smokers (because tobacco smokers were reported to have lower CSF CRH (Geracioti et al., 1997)), did not take any prescription or non-prescription medication and were healthy according to medical history, physical examination, standard blood tests and urinary drug screen. None had a history of shift work or transcontinental flights during the past 3 months. Current or life-time psychiatric disorders were excluded as per the Structured Clinical Interview for the DSM-IV (SCID), axis I. Subjects had no history of major psychological trauma according to the SCID post-traumatic stress disorder module's trauma screen. After full oral and written explanation of the purpose and procedures of the investigation, written informed consent was obtained from each subject. The study had been approved by the Ethical Committee of the Medical Board Hamburg.
663
120 100 80 60 40 20 0 11:00
12:00
13:00
Placebo
2.3. Side effects and complications No subject experienced acute side effects of the injection. Seven subjects developed postural headache in the late afternoon of the experimental day. While symptoms disappeared within a few days in two subjects, five were treated with an epidural blood patch, resulting in each case in rapid and sustained recovery. In one subject (from the cortisol treatment group) the catheter did not deliver CSF; therefore, hormone measurement was only possible in nine subjects. 2.4. Endocrine analyses CSF concentrations of cortisol were determined using a commercial radioimmunoassay (DRG, Marburg, Germany). Inter- and intraassay coefficients of variation were below 8%, detection limit was 0.5 ng/ml. CSF concentrations of CRH were directly measured with a radioimmunoassay as described earlier (Stalla et al., 1986). hCRH was used as a standard and N-tyr-hCRH as tracer after labeling with I-125. Intra- and interassay coefficients of variation were below 9%, lower limit of detection was 20 pg/ml. 2.5. Statistics The effect of cortisol vs. placebo on the variables CSF cortisol and CSF CRH was evaluated within the observational period via the curve indicators “mean location” (average of all time points) and “area under the curve without the linear background” (AUC). Multivariate analysis of variance was performed with treatment as within subjects factor. When a significant main effect was found, univariate F tests followed to identify the indicators which contributed significantly to this effect. In addition, polynomial contrasts for the group means at the sampling time points were calculated for CSF CRH in order to examine the course profiles over time in the treatment conditions. As nominal level of significance alpha ¼ 0.05 was accepted. All data are given as mean 7 standard deviation.
3. Results After intravenous cortisol injection, CSF cortisol rose steadily for about 90 min and reached a plateau with concentrations about 50-fold increased until the rest of the observational period (see Fig. 1). The mean locations were 65.67 11.0 ng/ml after cortisol and 2.4 7 1.6 ng/ml after placebo. Respective AUC values were 161.7 755.0 vs. 1.5 77.4.
cortisol
70
2.2. Procedure
60
CSF-CRH (pg/ml)
The subjects had fasted over-night and were studied from 08:00 to 14:00 (not sleeping, drinking or eating) in supine position in a sound-proof private room. At 08:00 an intravenous cannula was inserted into a forearm vein. Consecutively a subarachnoidal catheter (B. Braun AG, Melsungen, Germany) was placed in the lumbar 3–4 or 4–5 interspace by a skilled anesthesiologist after achieving adequate local anesthesia. At 11:02 an intravenous injection of 100 mg cortisol (Hydrocortison, Pfizer, Puurs, Belgium) or placebo (isovolemic normal saline) was given (in a single-blind manner and balancedly randomized). For the measurement of cortisol and CRH concentrations in CSF, from 10:40 to 14:00 every 20 min 2 ml were drawn from the subarachnoidal catheter (after discarding 0.5 ml because of the dead space in the conduit), placed immediately on ice into prechilled tubes and then stored at 80 1C until analysis.
14:00
50 40 30 20 10 0 11:00
12:00
13:00
14:00
Fig. 1. Cortisol and CRH concentrations in CSF (means7 standard deviations) in healthy men after an intravenous injection of 100 mg cortisol vs. placebo at 11:02 (arrow).
CSF CRH concentrations are shown in Fig. 1. While no effect of cortisol on them was obvious, levels increased slightly during the observational period in both treatment groups. The mean locations were 37.27 7.9 pg/ml after cortisol vs. 36.4 79.2 pg/ml after placebo. Respective AUC values were -23.0 7 31.7 vs. 2.6 754.2. Wilk's multivariate test of significance indicated significant treatment effects (F(4;4)¼ 27.26, sig of F¼0.004), located only to the CSF cortisol indicators mean location (F(1;7) ¼165.93, po 0.0001) and AUC (F(1;7) ¼ 44.63, po 0.0001). No significant treatment effects emerged for CSF CRH indicators mean location (F (1;7) ¼0.01, p ¼0.90) or AUC (F(1;7) ¼0.44, p ¼0.53). Polynomial contrasts for CSF CRH time concentration curves showed a significant linear trend both in the placebo (F(1;4) ¼ 13.55, p¼ 0.021) and the cortisol treatment condition (F(1;3) ¼ 13.33, p ¼0.035), pointing to a circadian increase of this peptide during our observational period.
4. Discussion We report first evidence that CSF CRH is not acutely suppressed by intravenous cortisol administration in healthy humans within 3 h, although cortisol in CSF was rapidly and considerably increased. Our results do not support that CSF CRH is a measure of hypothalamic CRH, but this has to be regarded as preliminary due to the small sample size and the time window in this pilot investigation. As Kling et al. (1994) have shown diurnal variation of CSF CRH in normal volunteers with concentrations highest in the evening and lowest in the morning (which has been replicated by D.G. Baker, personal communication), our initial hypothesis of an acute suppressive effect of cortisol on CSF CRH needs to be tested also in the evening. For such studies again young adult men should be studied, because they show the highest HPA axis sensitivity to cortisol feedback inhibition (Wilkinson et al., 1997).
664
M. Kellner et al. / Psychiatry Research 210 (2013) 662–664
As long as the precise anatomical sources of CRH in CSF, their relative contributions to CSF CRH concentrations under different circumstances and the physiological and pathophysiological role of CSF CRH have not sufficiently been elucidated, measurements of CRH in CSF seem difficult to interpret. In preclinical studies reduction of cortisol for several days by adrenalectomy resulted in differential effects on CRH-mRNA in various brain areas, e. g. a clear increase in the hypothalamus (Beyer et al., 1988; Frim et al., 1990; Imaki et al., 1991) and no change in the cortex (Frim et al., 1990; Imaki et al., 1991), while there was equivocal evidence for a decrease in the amygdala (Beyer et al., 1988; Palkovits et al., 1998). Thus, it may be hard to detect a clear-cut net signal of cortisol on CSF CRH concentration. Given a half-life time of CRH in CSF of about 10 min and because transfer from cephalad to lumbar spinal canal takes about 1 h in man (Geracioti et al., 1992), we cannot exclude having missed subtle short-term effects. However, because of a high correlation of lumbar and cisternal CRH concentrations, CSF sampled from lumbar regions is thought to adequately reflect changes at higher levels in the central nervous system (Kalin et al., 1987). According to the only study showing a significant transient reduction of CSF CRH in humans after an intervention (Kasckow et al., 2001), our observational period of 3 h might have been sufficient. On the other hand, in light of preclinical findings that after a stressful stimulus CRH mRNA reacted significantly not before 24 h (Imaki et al., 1991), CSF CRH levels should be tested also on the next day following cortisol administration. In addition, it cannot be excluded that sustained increases of cortisol levels might influence CSF CRH by delayed feedback effects. Further research is needed to determine the source and regulation of CSF CRH and its role in health and disease. Acknowledgment We thank Mrs. Kirsten Huwald, Mrs. Iris Remmlinger-Marten, and Mrs. Johanna Stalla for their expert technical help, Dr. Cornelius Fuchs for friendly advice and Dr. Martin Petzoldt for clinical cooperation. This manuscript is part of the M.D. thesis of TU. References Beyer, H.S., Matta, S.G., Sharp, B.M., 1988. Regulation of the messenger ribonucleic acid for corticotrophin-releasing factor in the paraventricular nucleus and other brain sites of the rat. Endocrinology 123, 2117–2123. deKloet, E.R., Joels, M., Holsboer, F., 2005. Stress and the brain: from adaptation to disease. Nature Reviews Neuroscience 6, 465–475.
Frim, D.M., Robinson, B.G., Pasieka, K.B., Majzoub, J.A., 1990. Differential regulation of corticotropin-relasing hormone mRNA in rat brain. American Journal of Physiology 258, E686–E692. Garrick, N.A., Hill, J.L., Szele, F.G., Tomai, T.P., Gold, P.W., Murphy, D.L., 1987. Corticotropin-releasing factor: a marked circadian rhythm in primate cerebrospinal fluid peaks in the evening an is inversely related to the cortisol circadian rhythm. Endocrinology 121, 1329–1334. Geracioti, T.D., Orth, D.N., Ekhator, N.N., Blumenkopf, B., Loosen, P.T., 1992. Serial cerebrospinal fluid corticotropin-releasing hormone concentrations in healthy and depressed humans. Journal of Clinical Endocrinology and Metabolism 74, 1325–1330. Geracioti, T.D., Loosen, P.T., Orth, D.N., 1997. Low cerebrospinal fluid corticotrophinreleasing hormone concentrations in eucortisolemic depression. Biological Psychiatry 42, 166–174. Hong, S.-K., Gold, P.W., Herkenham, M., 1993. Hypothalamic paraventricular nucleus lesions decrease corticotropin-releasing hormone in the CSF and elevate TH mRNA in the locus ceruleus. Society for Neuroscience Abstracts 19, 762. Imaki, T., Nahan, J.-L., Rivier, C., Sawchenko, P.E., Vale, W., 1991. Differential regulation of corticotrophin-releasing factor mRNA in rat brain regions by glucocorticoids and stress. The Journal of Neuroscience 11, 585–599. Kalin, N.H., Shelton, S.E., Barksdale, C.M., Brownfield, M.S., 1987. A diurnal rhythm in cerebrospinal fluid corticotrophin-releasing hormone different from the rhythm of pituitary-adrenal activity. Brain Research 426, 385–391. Kasckow, J.W., Hagan, M., Mulchahey, J.J., Baker, D.G., Ekhator, N.N., Strawn, J.R., Nicholson, W., Orth, D.N., Loosen, P.T., Geracioti, T.D., 2001. The effect of feeding on cerebrospinal fluid corticotropin-releasing hormone levels in humans. Brain Research 904, 218–224. Kling, M.A., DeBellis, M.D., O'Rourke, D.K., Listwak, S.J., Geracioti, T.D., McCutcheon, I.E., Kalogeras, K.T., Oldfield, E.H., Gold, P.W., 1994. Diurnal variation of cerebrospinal fluid immunoreactive corticotropin-releasing hormone levels in healthy volunteers. Journal of Clinical Endocrinology and Metabolism 79, 233–239. Lee, J.L., Hempel, J., TenHarmsel, A., Liu, T., Mathé, A.A., Klock, A., 2012. The neuroendocrinology of childhood trauma in personality disorder. Psychoneuroendocrinology 37, 78–86. Newport, D.J., Heim, C., Owens, M.J., Ritchie, J.C., Ramsey, C.H., Bonsall, R., Miller, A. H., Nemeroff, C.B., 2003. Cerebrospinal fluid corticotropin-releasing factor (CRF) and vasopressin concentrations predict pituitary response in the CRF stimulation test: a multiple regression analysis. Neuropsychopharmacology 28, 569–576. Palkovits, M., Young, W.S., Kovács, K., Tóth, Z., Makara, G.B., 1998. Alterations in corticotropin-releasing hormone gene expression of central amydaloid neurons following long-term paraventricular lesions and adrenalectomy. Neuroscience 85, 135–147. Rock, J.P., Oldfield, E., Schulte, H.M., Gold, P.W., Kornblith, P.L., Loriaux, L., Chrousos, G.P., 1984. Corticotropin releasing factor administered into the ventricular CSF stimulates the pituitary–adrenal axis. Brain Research 323, 365–368. Stalla, G.K., Hartwimmer, J., Schopohl, J., von Werder, K., Müller, A.O., 1986. Intravenous application of ovine and human corticotropin releasing factor (CRF): ACTH, cortisol and CRF levels. Neuroendocrinology 42, 1–5. Vythilingam, M., Anderson, G.M., Owens, M.J., Halaszynski, T.M., Bremner, J.D., Carpenter, L.L., Henninger, G.R., Nemeroff, D.B., Charney, D.S., 2000. Cerebrospinal fluid corticotropin-releasing hormone in healthy humans: effects of yohimbine and naloxone. Journal of Clinical Endocrinology and Metabolism 85, 4138–4145. Wilkinson, C.W., Peskind, E.R., Raskind, M.A., 1997. Decreased hypothalamicpituitary-adrenal axis sensitivity to cortisol feedback inhibition in human aging. Neuroendocrinology 65, 79–90.
Contents lists available at ScienceDirect
Psychiatry Research journal homepage: www.elsevier.com/locate/psychres
Brief report
No acute suppression of cerebrospinal fluid corticotropin-releasing hormone in man by cortisol administration Michael Kellner a,n, Cornelie Salzwedel b, Viola Wortmann a, Tatiana Urbanowicz a, Kai Boelmans a, Alexander Yassouridis c, Günter K. Stalla c, Klaus Wiedemann a a
University Hospital Hamburg-Eppendorf, Department of Psychiatry and Psychotherapy, Hamburg, Germany University Hospital Hamburg-Eppendorf, Department of Anesthesiology, Hamburg, Germany c Max Planck Institute of Psychiatry, Munich, Germany b
art ic l e i nf o
a b s t r a c t
Article history: Received 9 January 2013 Received in revised form 18 June 2013 Accepted 7 July 2013
Corticotropin-releasing hormone (CRH) in cerebrospinal fluid (CSF) is regarded as index of brain endocrine and behavioral functioning. We investigated the acute effects of intravenous cortisol (100 mg) vs. placebo on serial CSF CRH in ten healthy men. CSF CRH concentrations were not significantly suppressed by cortisol within 3 h. The origin and regulation of CSF CRH need further research. & 2013 Elsevier Ireland Ltd. All rights reserved.
Keywords: Corticotropin-releasing hormone (CRH) Cortisol Cerebrospinal fluid (CSF)
1. Introduction Corticotropin-releasing hormone (CRH) has a major role in stress response. CRH neurons from the hypothalamic paraventricular nucleus (PVN) control hypothalamic-pituitary-adrenocortical (HPA) activity. Furthermore, extra-hypothalamic CRH has been found in the brain in cortical, limbic (central nucleus of the amygdala, bed nucleus of the stria terminalis, hippocampus) and brainstem regions, where it modulates autonomic and behavioral functions, such as anxiety, food intake, sleep, arousal, locomotion, learning and memory. Therefore, CRH and HPA system dysfunctions have been hypothesized to be implicated in the pathophysiology of psychiatric diseases, particularly of mood and anxiety disorders (deKloet et al., 2005). CRH concentrations have been measured in cerebrospinal fluid (CSF) of human subjects as a putative index of central nervous system CRH functioning (Geracioti et al., 1992). However, the source and regulation of CSF CRH have not been sufficiently clarified. Following bilateral ablation of the PVN, CSF CRH declined in rats suggesting that a considerable portion may be hypothalamic (Hong et al., 1993). In monkeys intracerebroventricular CRH stimulated the pituitary–adrenocortical axis (Rock et al., 1984).
n Correspondence to: University Hospital Hamburg-Eppendorf, Dept. of Psychiatry and Psychotherapy, Martinistrasse 52, W37, D-20246 Hamburg, Germany. Tel.: +49 40 7410 52234; fax: +49 40 7410 53461. E-mail address: [email protected] (M. Kellner).
0165-1781/$ - see front matter & 2013 Elsevier Ireland Ltd. All rights reserved. http://dx.doi.org/10.1016/j.psychres.2013.07.003
However, the diurnal rhythms of CSF CRH and of plasma pituitaryadrenocortical hormones in non-human primates were not closely linked (Garrick et al., 1987; Kalin et al., 1987); despite HPA activation by physiostigmine, metyrapone (Kalin et al., 1987) and m-chlorophenylpiperazine (Garrick et al., 1987) no consecutive changes in CSF CRH were observed. Thus, CSF CRH may rather reflect extra-hypothalamic than hypophysiotropic brain functions of this peptide. Also in man lack of synchrony between lumbar CSF CRH concentrations and pituitary–adrenocortical activity was demonstrated in normal volunteers and depressed patients (Geracioti et al., 1992, 1997). In contrast, Kling et al. (1994) reported significant negative correlations of CSF CRH and simultaneous plasma cortisol in man and suggested partial glucocorticoid suppressibility of CRH secretion into CSF. Complementarily, both Newport et al. (2003) and Lee et al. (2012) found that the CRH concentration in human CSF was inversely correlated with pituitary responsiveness to exogenous CRH. Kasckow et al. (2001) observed that after feeding CSF CRH diminished in man, which may be caused by rises in plasma cortisol and associated negative feedback effects. On the other hand, Vythilingam et al. (2000) reported that the hypothalamic CRH-releaser naloxone increased plasma cortisol, but no concomitant change of CSF CRH was seen in healthy humans. Interestingly, so far the acute effects of cortisol administration on CSF CRH have not been investigated. Therefore, we performed a respective pilot study in healthy human volunteers using a serial CSF sampling technique and hypothesized suppressive effects of cortisol on CSF CRH.
M. Kellner et al. / Psychiatry Research 210 (2013) 662–664
Placebo
2. Methods
cortisol
140
CSF-Cortisol (ng/ml)
2.1. Participants Ten young men (mean age 22.7 years, range 21–27; mean body mass index 24.3 kg/m2, range 22.3–27.5) were studied. They all were non-smokers (because tobacco smokers were reported to have lower CSF CRH (Geracioti et al., 1997)), did not take any prescription or non-prescription medication and were healthy according to medical history, physical examination, standard blood tests and urinary drug screen. None had a history of shift work or transcontinental flights during the past 3 months. Current or life-time psychiatric disorders were excluded as per the Structured Clinical Interview for the DSM-IV (SCID), axis I. Subjects had no history of major psychological trauma according to the SCID post-traumatic stress disorder module's trauma screen. After full oral and written explanation of the purpose and procedures of the investigation, written informed consent was obtained from each subject. The study had been approved by the Ethical Committee of the Medical Board Hamburg.
663
120 100 80 60 40 20 0 11:00
12:00
13:00
Placebo
2.3. Side effects and complications No subject experienced acute side effects of the injection. Seven subjects developed postural headache in the late afternoon of the experimental day. While symptoms disappeared within a few days in two subjects, five were treated with an epidural blood patch, resulting in each case in rapid and sustained recovery. In one subject (from the cortisol treatment group) the catheter did not deliver CSF; therefore, hormone measurement was only possible in nine subjects. 2.4. Endocrine analyses CSF concentrations of cortisol were determined using a commercial radioimmunoassay (DRG, Marburg, Germany). Inter- and intraassay coefficients of variation were below 8%, detection limit was 0.5 ng/ml. CSF concentrations of CRH were directly measured with a radioimmunoassay as described earlier (Stalla et al., 1986). hCRH was used as a standard and N-tyr-hCRH as tracer after labeling with I-125. Intra- and interassay coefficients of variation were below 9%, lower limit of detection was 20 pg/ml. 2.5. Statistics The effect of cortisol vs. placebo on the variables CSF cortisol and CSF CRH was evaluated within the observational period via the curve indicators “mean location” (average of all time points) and “area under the curve without the linear background” (AUC). Multivariate analysis of variance was performed with treatment as within subjects factor. When a significant main effect was found, univariate F tests followed to identify the indicators which contributed significantly to this effect. In addition, polynomial contrasts for the group means at the sampling time points were calculated for CSF CRH in order to examine the course profiles over time in the treatment conditions. As nominal level of significance alpha ¼ 0.05 was accepted. All data are given as mean 7 standard deviation.
3. Results After intravenous cortisol injection, CSF cortisol rose steadily for about 90 min and reached a plateau with concentrations about 50-fold increased until the rest of the observational period (see Fig. 1). The mean locations were 65.67 11.0 ng/ml after cortisol and 2.4 7 1.6 ng/ml after placebo. Respective AUC values were 161.7 755.0 vs. 1.5 77.4.
cortisol
70
2.2. Procedure
60
CSF-CRH (pg/ml)
The subjects had fasted over-night and were studied from 08:00 to 14:00 (not sleeping, drinking or eating) in supine position in a sound-proof private room. At 08:00 an intravenous cannula was inserted into a forearm vein. Consecutively a subarachnoidal catheter (B. Braun AG, Melsungen, Germany) was placed in the lumbar 3–4 or 4–5 interspace by a skilled anesthesiologist after achieving adequate local anesthesia. At 11:02 an intravenous injection of 100 mg cortisol (Hydrocortison, Pfizer, Puurs, Belgium) or placebo (isovolemic normal saline) was given (in a single-blind manner and balancedly randomized). For the measurement of cortisol and CRH concentrations in CSF, from 10:40 to 14:00 every 20 min 2 ml were drawn from the subarachnoidal catheter (after discarding 0.5 ml because of the dead space in the conduit), placed immediately on ice into prechilled tubes and then stored at 80 1C until analysis.
14:00
50 40 30 20 10 0 11:00
12:00
13:00
14:00
Fig. 1. Cortisol and CRH concentrations in CSF (means7 standard deviations) in healthy men after an intravenous injection of 100 mg cortisol vs. placebo at 11:02 (arrow).
CSF CRH concentrations are shown in Fig. 1. While no effect of cortisol on them was obvious, levels increased slightly during the observational period in both treatment groups. The mean locations were 37.27 7.9 pg/ml after cortisol vs. 36.4 79.2 pg/ml after placebo. Respective AUC values were -23.0 7 31.7 vs. 2.6 754.2. Wilk's multivariate test of significance indicated significant treatment effects (F(4;4)¼ 27.26, sig of F¼0.004), located only to the CSF cortisol indicators mean location (F(1;7) ¼165.93, po 0.0001) and AUC (F(1;7) ¼ 44.63, po 0.0001). No significant treatment effects emerged for CSF CRH indicators mean location (F (1;7) ¼0.01, p ¼0.90) or AUC (F(1;7) ¼0.44, p ¼0.53). Polynomial contrasts for CSF CRH time concentration curves showed a significant linear trend both in the placebo (F(1;4) ¼ 13.55, p¼ 0.021) and the cortisol treatment condition (F(1;3) ¼ 13.33, p ¼0.035), pointing to a circadian increase of this peptide during our observational period.
4. Discussion We report first evidence that CSF CRH is not acutely suppressed by intravenous cortisol administration in healthy humans within 3 h, although cortisol in CSF was rapidly and considerably increased. Our results do not support that CSF CRH is a measure of hypothalamic CRH, but this has to be regarded as preliminary due to the small sample size and the time window in this pilot investigation. As Kling et al. (1994) have shown diurnal variation of CSF CRH in normal volunteers with concentrations highest in the evening and lowest in the morning (which has been replicated by D.G. Baker, personal communication), our initial hypothesis of an acute suppressive effect of cortisol on CSF CRH needs to be tested also in the evening. For such studies again young adult men should be studied, because they show the highest HPA axis sensitivity to cortisol feedback inhibition (Wilkinson et al., 1997).
664
M. Kellner et al. / Psychiatry Research 210 (2013) 662–664
As long as the precise anatomical sources of CRH in CSF, their relative contributions to CSF CRH concentrations under different circumstances and the physiological and pathophysiological role of CSF CRH have not sufficiently been elucidated, measurements of CRH in CSF seem difficult to interpret. In preclinical studies reduction of cortisol for several days by adrenalectomy resulted in differential effects on CRH-mRNA in various brain areas, e. g. a clear increase in the hypothalamus (Beyer et al., 1988; Frim et al., 1990; Imaki et al., 1991) and no change in the cortex (Frim et al., 1990; Imaki et al., 1991), while there was equivocal evidence for a decrease in the amygdala (Beyer et al., 1988; Palkovits et al., 1998). Thus, it may be hard to detect a clear-cut net signal of cortisol on CSF CRH concentration. Given a half-life time of CRH in CSF of about 10 min and because transfer from cephalad to lumbar spinal canal takes about 1 h in man (Geracioti et al., 1992), we cannot exclude having missed subtle short-term effects. However, because of a high correlation of lumbar and cisternal CRH concentrations, CSF sampled from lumbar regions is thought to adequately reflect changes at higher levels in the central nervous system (Kalin et al., 1987). According to the only study showing a significant transient reduction of CSF CRH in humans after an intervention (Kasckow et al., 2001), our observational period of 3 h might have been sufficient. On the other hand, in light of preclinical findings that after a stressful stimulus CRH mRNA reacted significantly not before 24 h (Imaki et al., 1991), CSF CRH levels should be tested also on the next day following cortisol administration. In addition, it cannot be excluded that sustained increases of cortisol levels might influence CSF CRH by delayed feedback effects. Further research is needed to determine the source and regulation of CSF CRH and its role in health and disease. Acknowledgment We thank Mrs. Kirsten Huwald, Mrs. Iris Remmlinger-Marten, and Mrs. Johanna Stalla for their expert technical help, Dr. Cornelius Fuchs for friendly advice and Dr. Martin Petzoldt for clinical cooperation. This manuscript is part of the M.D. thesis of TU. References Beyer, H.S., Matta, S.G., Sharp, B.M., 1988. Regulation of the messenger ribonucleic acid for corticotrophin-releasing factor in the paraventricular nucleus and other brain sites of the rat. Endocrinology 123, 2117–2123. deKloet, E.R., Joels, M., Holsboer, F., 2005. Stress and the brain: from adaptation to disease. Nature Reviews Neuroscience 6, 465–475.
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