Hypothalamic changes during the immune response
Eur. J. Immunol. 1977.7: 323-325
323
Short Papers H. Besedovsky+, E. Sorkin+, D. Felix' and H. Haas' Schweizerisches Fonchungsinstitut, Medirinische Abteilung+, Davos, Hirnfonchungsinstitutder Univenitiit' and NeurochirurgischaUnivenitiitsklinik', Zurich
Hypothalamic changes during the immune response The immune system is subject to an array of identified autoregulatory processes, but immunoregulation may also have a further basis in a network of immune-neuroendocrine interactions. Two antigens each produced an increase of more than 100 % in electrical activity of individual neurones in the ventromedial but not in the anterior nucleus of the rat hypothalamus. Animals that failed to respond to antigen manifested no increase in the firing rate. These findings constitute the first evidence for a flow of information from the activated immune system to the hypothalamus, suggesting that the brain is involved in the immune response.
1. Introduction
2.2. Recording of hypothalamic activity
The immune system is subject t o self-monitoring and selfregulating processes which are still incompletely understood [ 11. In view of its critical importance in coping with threats of the environment this system is unlikely to function in a completely autonomous manner. As with most other systems it is presumably subject t o regulation from the nervous and the endocrine systems. We have recently noted that after antigenic stimulation of rats there was a striking increase in the serum levels of corticosterone and a moderate decrease in thyroxine, coinciding in time with the elaboration of antibody-forming cells [2-41. We regard this as an expression of the response of neuroendocrine structures t o signals deriving originally from activated lymphoid cells. In order to obtain direct evidence for such a proposed afferent pathway, we looked for physiological changes manifested as electrical activity of individual neurones in the ventromedial nucleus of the hypothalamus of rats after immunization with two different antigens. This site is known t o be involved in many autonomous and endocrine control mechanisms.
For measuring the rate of firing of unitary neurones in the ventromedial part of the hypothalamus the animals were anesthetized i.p. with 1350 mglkg-' urethane. The animals were fixed in a HorselyClarke stereotactic apparatus, the electrode placed into the ventromedial nucleus. Action potentials were recorded extracellularly with monopolar platinum-iridium electrodes (tip diameter 1 flm, resistance 3 Ma), or with glass micropipettes filled with 4 M NaCl or pontamine sky blue (2 % in 0.5 M sodium acetate). In several animals histological examination of electrode tracks of dye marks ejected from the electrode, confirmed the location of the recording sites. The recording electrodes were connected t o a negative capacitance cathode follower. The spontaneous activity was displayed on an oscilloscope and plotted directly on a UV oscillograph. The rate of firing on each neurone was counted by a ratemeter and displayed continuously on the UV oscillograph. The discharge of each neurone was tested over a period of 200 sec. In addition t o the chart recording, the mean frequency was enumerated by means of a programmable countertimer.
2. Materials and methods 2.1. Animals and antigens
A total of 98 female Holtzmann rats (Tierfarm Ftillinsdorf, Switzerland) were used in this study. 49 rats were injected with antigen. Groups were immunized intraperitoneally with 5 x lo9 sheep red blood cells (SRBC) or 250 pg trinitrophenylated hemocyanin (TNP-HE) [5]. 44 rats received control injections of 1 ml of diluent (0.9 % NaCl) i.p. The animals were kept in individual cages at least one week before injection and until completion of the experiments. The kinetics of the immune response t o these antigens was determined in numerous Holtzmann rats, using the Jerne technique for plaque formation in the spleen. Direct plaque formation to SRBC occurred from day 4 t o day 7 with a peak at days 5-6 and to TNP-HE on day 1 to day 4 with a peak on day 3. [I 16101 * This work was supported by the Swiss National Science Foundation, Grant Nr. 3.600.75 (H.B. and E.S.) and Nr. 3.534.0.75 (D.F.). ~~~
Correspondence: Ernst Sorkin, Schweizerisches Forschungsinstitut, Medizinische Abteilung, CH-7270 Davos, Switzerland Abbreviations: SRBC: Sheep red blood cells TNP-HE:Trinitrophenylated hemocyanin PFC: Plaque-forming cell(s)
~~
A total of 490 individual neurones were examined. Control and immunized animals were recorded alternately. After recording, the level of antibody formation in individual rats immunized with SRBC was also determined.
3. Results 3.1. Hypothalamic response after stimulation with
SRBC
Typical data for 14 of 29 animals stimulated with SRBC are summarized in Fig. 1 a and b. On day 1 when no plaqueforming cells (PFC) are evident, no changes in firing frequency were demonstrable compared with controls. On day 5 , at or near peak of PFC in spleen, there was a more than twofold increase in the firing rate of the ventromedial neurones (Control: 3.75 f 0.28 spikes/sec.; SRBC: 8.82 1.30 spikeslsec., p < 0.001). In several rats which were immunological nonresponders, no increase in firing rates occurred (not shown in Fig. 1). Furthermore, no changes in firing rates were observed in the anterior hypothalamic nucleus (Control: 5.77 f 1 . 1 1 ;SRBC: 5.07 f 0.96) on day 5 after SRBC injection.
*
Eur. J. Immunol. 1977. 7: 323-325
H. Besedovsky, E. Sorkin, D. Felix and H. Haas
324
PFC (Control: 3.23 f 0.36 spikes/sec; TNP-HE: 6.84 f 0.78 spikes/sec., p < 0.001). The reason why there is n o change in firing rate o n day 3 cannot be explained.
1
3.3. Hypothalamic response following wound healing and adrenalectomy
SRBC IDAV 11 7 RATS
The corticosteroid blood levels increase during the immune response [2], and this increase might be one of the causes of the observed changes of firing rates in the ventromedial hypothalamic nucleus. In order t o exclude this possibility, 8 rats were adrenalectomized and controls sham-operated. The firing rates of neurones of the ventromedial part of the hypothalamus were measured 5- 10 days after surgery. The following results were obtained: adrenalectomized: 4.03 5 0.62; sham-operated controls: 2.86 f 0.41. It is concluded that the described increase in firing rates produced by the antigens can hardly be related to the increased corticosteroid levels.
SRBC IDAV 51 7RATS T
ma
IOAV 51 42 NEWONES 1 RATS P
Eur. J. Immunol. 1977.7: 323-325
323
Short Papers H. Besedovsky+, E. Sorkin+, D. Felix' and H. Haas' Schweizerisches Fonchungsinstitut, Medirinische Abteilung+, Davos, Hirnfonchungsinstitutder Univenitiit' and NeurochirurgischaUnivenitiitsklinik', Zurich
Hypothalamic changes during the immune response The immune system is subject to an array of identified autoregulatory processes, but immunoregulation may also have a further basis in a network of immune-neuroendocrine interactions. Two antigens each produced an increase of more than 100 % in electrical activity of individual neurones in the ventromedial but not in the anterior nucleus of the rat hypothalamus. Animals that failed to respond to antigen manifested no increase in the firing rate. These findings constitute the first evidence for a flow of information from the activated immune system to the hypothalamus, suggesting that the brain is involved in the immune response.
1. Introduction
2.2. Recording of hypothalamic activity
The immune system is subject t o self-monitoring and selfregulating processes which are still incompletely understood [ 11. In view of its critical importance in coping with threats of the environment this system is unlikely to function in a completely autonomous manner. As with most other systems it is presumably subject t o regulation from the nervous and the endocrine systems. We have recently noted that after antigenic stimulation of rats there was a striking increase in the serum levels of corticosterone and a moderate decrease in thyroxine, coinciding in time with the elaboration of antibody-forming cells [2-41. We regard this as an expression of the response of neuroendocrine structures t o signals deriving originally from activated lymphoid cells. In order to obtain direct evidence for such a proposed afferent pathway, we looked for physiological changes manifested as electrical activity of individual neurones in the ventromedial nucleus of the hypothalamus of rats after immunization with two different antigens. This site is known t o be involved in many autonomous and endocrine control mechanisms.
For measuring the rate of firing of unitary neurones in the ventromedial part of the hypothalamus the animals were anesthetized i.p. with 1350 mglkg-' urethane. The animals were fixed in a HorselyClarke stereotactic apparatus, the electrode placed into the ventromedial nucleus. Action potentials were recorded extracellularly with monopolar platinum-iridium electrodes (tip diameter 1 flm, resistance 3 Ma), or with glass micropipettes filled with 4 M NaCl or pontamine sky blue (2 % in 0.5 M sodium acetate). In several animals histological examination of electrode tracks of dye marks ejected from the electrode, confirmed the location of the recording sites. The recording electrodes were connected t o a negative capacitance cathode follower. The spontaneous activity was displayed on an oscilloscope and plotted directly on a UV oscillograph. The rate of firing on each neurone was counted by a ratemeter and displayed continuously on the UV oscillograph. The discharge of each neurone was tested over a period of 200 sec. In addition t o the chart recording, the mean frequency was enumerated by means of a programmable countertimer.
2. Materials and methods 2.1. Animals and antigens
A total of 98 female Holtzmann rats (Tierfarm Ftillinsdorf, Switzerland) were used in this study. 49 rats were injected with antigen. Groups were immunized intraperitoneally with 5 x lo9 sheep red blood cells (SRBC) or 250 pg trinitrophenylated hemocyanin (TNP-HE) [5]. 44 rats received control injections of 1 ml of diluent (0.9 % NaCl) i.p. The animals were kept in individual cages at least one week before injection and until completion of the experiments. The kinetics of the immune response t o these antigens was determined in numerous Holtzmann rats, using the Jerne technique for plaque formation in the spleen. Direct plaque formation to SRBC occurred from day 4 t o day 7 with a peak at days 5-6 and to TNP-HE on day 1 to day 4 with a peak on day 3. [I 16101 * This work was supported by the Swiss National Science Foundation, Grant Nr. 3.600.75 (H.B. and E.S.) and Nr. 3.534.0.75 (D.F.). ~~~
Correspondence: Ernst Sorkin, Schweizerisches Forschungsinstitut, Medizinische Abteilung, CH-7270 Davos, Switzerland Abbreviations: SRBC: Sheep red blood cells TNP-HE:Trinitrophenylated hemocyanin PFC: Plaque-forming cell(s)
~~
A total of 490 individual neurones were examined. Control and immunized animals were recorded alternately. After recording, the level of antibody formation in individual rats immunized with SRBC was also determined.
3. Results 3.1. Hypothalamic response after stimulation with
SRBC
Typical data for 14 of 29 animals stimulated with SRBC are summarized in Fig. 1 a and b. On day 1 when no plaqueforming cells (PFC) are evident, no changes in firing frequency were demonstrable compared with controls. On day 5 , at or near peak of PFC in spleen, there was a more than twofold increase in the firing rate of the ventromedial neurones (Control: 3.75 f 0.28 spikes/sec.; SRBC: 8.82 1.30 spikeslsec., p < 0.001). In several rats which were immunological nonresponders, no increase in firing rates occurred (not shown in Fig. 1). Furthermore, no changes in firing rates were observed in the anterior hypothalamic nucleus (Control: 5.77 f 1 . 1 1 ;SRBC: 5.07 f 0.96) on day 5 after SRBC injection.
*
Eur. J. Immunol. 1977. 7: 323-325
H. Besedovsky, E. Sorkin, D. Felix and H. Haas
324
PFC (Control: 3.23 f 0.36 spikes/sec; TNP-HE: 6.84 f 0.78 spikes/sec., p < 0.001). The reason why there is n o change in firing rate o n day 3 cannot be explained.
1
3.3. Hypothalamic response following wound healing and adrenalectomy
SRBC IDAV 11 7 RATS
The corticosteroid blood levels increase during the immune response [2], and this increase might be one of the causes of the observed changes of firing rates in the ventromedial hypothalamic nucleus. In order t o exclude this possibility, 8 rats were adrenalectomized and controls sham-operated. The firing rates of neurones of the ventromedial part of the hypothalamus were measured 5- 10 days after surgery. The following results were obtained: adrenalectomized: 4.03 5 0.62; sham-operated controls: 2.86 f 0.41. It is concluded that the described increase in firing rates produced by the antigens can hardly be related to the increased corticosteroid levels.
SRBC IDAV 51 7RATS T
ma
IOAV 51 42 NEWONES 1 RATS P
