Acta Physiologica Sinica465

Dec. 2001, 53 (6), 465¡«468

The role of central arginine vasopressin in corticotropin releasing hormoneª²induced fever in rats

WANG Huaª²Dong1,*, WANG Yanª²Ping1, HU Caoª²Feng1, QI Renª²Bin1, YAN Yuª²Xia2, LU Daª²Xiang1, LI Chuª²Jie1,

(Departments of ~1Pathophysiology and ~2Biochemistry, Medical College of Jinan University, Guangzhou 510632)

Abstract: The purpose of the present study was to investigate the role of central arginine vasopressin (AVP) in corticotropin releasing hormone (CRH)ª²induced fever in the rat. Guide cannulae were inserted into the third ventricle and placed over the ventral septal area (VSA). The content of arginine vasopressin in the VSA of the brain was determined by radioimmunoassay. Colon temperature was monitored in lightly restrained rats by insertion of a catheterª²mounted thermistor probe 5 cm in the rectum. The results demonstrated that intracerebroventricular (icv) injection of CRH increased AVP level in the VSA and the colonic temperature of the rats. Microinjection of AVP V1 antagonist into the VSA 10 min before CRH administration significantly enhanced CRHª²induced febrile response, while AVP V1 antagonist itself did not have a significant effect on the colonic temperature. Furthermore, injection of AVP into the VSA 5 min before CRH administration (icv) suppressed the fever evoked by CRH. These findings suggest that CRH is an important factor that stimulates the release of AVP in the VSA during fever, and endogenous AVP in the VSA has an antipyretic action on the CRHª²induced fever.

Key words: corticotrophin releasing hormone; arginine vasopressin; fever; thermoregulation

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Íõ»ª¶«1,*, ÍõÑåƽ1, ºú³²·ï1, ÆÝÈʱó1, ÑÏÓñϼ2, ½´óÏé1, Àî³þ½Ü1

(1ôßÄÏ´óѧҽѧԺ²¡ÀíÉúÀí½ÌÑÐÊÒ, 2Éú»¯½ÌÑÐÊÒ, ¹ãÖÝ 510632)

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ѧ¿Æ·ÖÀàºÅ: Q426; R364.6

It has been demonstrated that central corticotrophin releasing hormone (CRH) plays an important role in the febrile response in addition to its major actions on the hypothalamicª²pituitaryª²adrenal axis. Intracerebroventricular injection of CRH causes a rapid increase in body temperature, and the pyrogenic actions of interleukinª²1¦Â (ILª²1¦Â), ILª²6, ILª²8, and lipopolysaccharide can all be prevented by inhibiting CRH action with antiª²CRH polyclonal or monoclonal antibodies or with CRH receptor antagonist£Û1¡«3£Ý, which suggests that CRH is a central pyrogenic mediator of fever. However, some studies have controversial evidence that central administration of CRH has an antipyretic action£Û4,5£Ý. Therefore, the mechanism underlying thermoregulatory action of CRH is to be further elucidated.

On the other hand, now there is evidence that arginine vasopressin (AVP) acts as a neurotransmitter in the ventral septal area (VSA) to reduce fever. Dialysate of the VSA shows that increased AVP levels during defervescence, and exogenous administration of AVP in the VSA of a variety of species results in an attenuation of fever without performing an effect on afebrile body temperature. Centrally injected AVP V1ª²type receptor antagonist or AVP antiserum causes prolonged and exacerbated fever£Û6,7£Ý. It is possible that central thermoregulatory action of CRH involves the release of AVP in the VSA. The aim of the present study was to test this hypothesis by investigating the effects of centrally administered CRH on the colonic temperature and AVP level in the VSA, and by examining the effects of injection of AVP and AVP V1 antagonist into the VSA on the CRHª²induced febrile response.

1MATERIALS AND METHODS

1.1 AnimalsAdult male Spragueª²Dawley rats (200¡«250 g, obtained from the Experimental Animal Center, Sun Yatª²Sen University of Medical Sciences; grade ¢ò, certificate 98A066) were used for the experiments. Animals were caged individually and had free access to food and water. To minimize the stress response caused by handling during experimentation, animals were confined in cages at room temperature of (22¡À1)¡æ. At the same time, a catheter mimicking the thermistor probe was inserted 5 cm into the colon of the rat and fixed on the root of the tail for 2 h/d for 3 d. All experimental trials were carried out between 08¡Ã00 and 16¡Ã00, the time when the body temperature in the rat is relatively stable according to the observation by Federico et al£Û8£Ý.

1.2 Surgical ProceduresUnder sodium pentobarbital anesthesia (60 mg/kg, ip), a guide cannula was inserted into the third ventricle as described previously£Û9£Ý, and secured to the skull using dental cement. Some of the rats were also implanted with bilateral guide cannula placed over the VSA (coordinates: bregma +2.8 mm; midline ¡À1.0 mm; dura £3.0 mm)£Û10£Ý. The animals were allowed to recover for at least 5 days before experimentation. Positioning of cannula was verified histologically at the end of the experiment.

1.3 Body temperature response measurementDuring experiments each rat was placed individually in a plastic cage without restraint. Colon temperature was monitored in lightly restrained rats by insertion of a catheterª²mounted thermistor probe 5 cm in the rectum, the thermistors were fed into a telethermometer (STª²1 Type, Shanghai Medical Instruments Factory). Temperature records were taken at 30 min intervals until the experiment was over. The febrile response was represented by the thermal response index (TRI) which was calculated by integration of the area under the mean thermal response curve.

1.4 Administration of test substancesRat CRH, AVP and the AVP V1 receptor antagonist, d (CH2)5Tyr (Me)AVP were purchased from Sigma Chemicals Ltd and dissolved in 0.9% sterile saline, which was employed as a vehicle control. CRH or normal saline (5 ¦Ìl) was injected into the third ventricle via the guide cannula in lightly restrained rats over a period of 1¡«2 min. Ten min before CRH administration, either normal saline (1.0 ¦Ìl) or AVP V1 receptor antagonist (1.0 ¦Ìl) was injected bilaterally into the VSA over 30¡«60 s. AVP (1.0 ¦Ìl) was injected bilaterally into the VSA 5 min before injection of CRH.

1.5 Determination of AVP contentThe rats were decapitated at the end of the experiments, the brain was removed quickly and boiled in normal saline for 30 min. The VSA was dissected for AVP assay. The concentration of AVP in the VSA was determined by radioimmunoassay as described earlier using AVP assay kits (obtained from the Second Military Medical University, Shanghai, China)£Û12£Ý. Protein content was measured by the method of Lowry et al£Û13£Ý.

1.6 Procedures for prevention of contaminationThe laboratory and all materials were routinely sterilized using the method previously described£Û11£Ý, experiments were performed under aseptic conditions.

1.7 Statistical analysisThe data are given as the mean¡ÀSD. Results were analyzed using oneª²way analysis of variance procedures followed by the Studentª²Newmenª²Keul¡äs test for multiple comparisons of means. Student¡äs t test was used for comparison of the two means. Differences were considered to be significant if P<0.05.

2RESULTS

2.1 The colonic temperature response to intracerebroventricular injection of CRH

Effects of injection (icv) of 5 ¦Ìl 0.9% sterile saline (NS) and 5 ¦Ìl CRH (5 ¦Ìg) on changes in the colonic temperature of rats were examined. Changes in the colonic temperature are expressed as a deviation from the baseline recorded at the start time of the experiment. The ranges of colonic temperature in the NS group were less than 0.3¡æ. Injection (icv) of CRH at a dose of 5 ¦Ìg caused a rapid rise in the colonic temperature. (Fig.1, Table 1)

Fig.1.Mean colonic temperature responses evoked by an intracerebroventricular injection of CRH (5.0 ¦Ìg). Mean¡ÀSD. ~*P<0.05 vs NS group.

Table¡¡£±.Effect of CRH (5.0 ¦Ìg) on TRI2 and AVP content in the VSA in rats (mean¡ÀSD)

**~**P<0.05 vs NS group. NS, normal saline. TRI2, 2 h thermal response index.

2.2 Effect of injection (icv) of CRH on AVP content in the VSA

As shown in Table 1,¡¡intracerebroventricular administration of CRH (5 ¦Ìg) caused a significant increase in AVP level of the VSA compared with control group.

2.3 Effect of microinjection of AVP V1 antagonist into VSA on the CRHª²induced febrile response

¡¡Injection of either saline or the AVP V1 antagonist into the VSA, in the absence of CRH, did not evoke any consistent alteration in the colonic temperature of the rats. When the icv injection of CRH (2.5 ¦Ìg) was preceded by a bilateral injection of normal saline into the VSA, fever was induced which was not different from that produced by CRH alone (data not shown). However, when the AVP V1 antagonist (3 ¦Ìg) was injected bilaterally into the VSA 10 min prior to administration of CRH (2.5 ¦Ìg) into the third ventricle, the 2 h thermal response indexes of the rats were significant greater than those obtained from the rats that were treated with normal saline (VSA) and CRH (2.5 ¦Ìg, icv)¡¡(Fig.2, Table 2)£®

Fig.2. Effect of AVP V1 receptor antagonist on CRHª²induced febrile response in conscious rats. AVP V1 receptor antagonist (V1A, 3 ¦Ìg) or saline (NS) was injected into the VSA 10 min prior to CRH (2.5 ¦Ìg, icv) or saline at time 0. mean¡ÀSD. ~*P<0.05 vs NS+NS group, ~#P<0.05 vs NS+CRH group.

Table 2. 2 h thermal response index (TRI2) in various groups shown in Fig.2 (mean¡ÀSD)

~*P>0.05, ~**P<0.05 vs NS+NS group. ~#P<0.05 vs NS+CRH group.

2.4 Effect of AVP pretreatment on CRHª²induced fever

As shown in Table 3, treatment of AVP (60 pmol) 5 min before CRH significantly suppressed the febrile response to administration of CRH (5 ¦Ìg, icv) in the rat.

Table¡¡3.Effect of injection of AVP into the VSA on CRHª²induced febrile response in rats (mean¡ÀSD)

~#P<0.01 vs NS+NS group, ~*P<0.05 vs NS+CRH group. NS, normal saline. TRI2, 2 h thermal response index.

3DISCUSSION

¡¡Some studies have demonstrated that central CRH mediates febrile response induced by some pyrogenic cytokines, the role of CRH is dependent on central release of proª²opiomelanocortin and sympathetic activation of brown adipose tissue£Û1£Ý, which suggests that central CRH itself may be an important mediator of fever. We also observed that the pyrogenic effect of centrally administration of CRH on the rat was mediated by the cAMPª²dependent protein kinase signal transduction pathway (data not shown). However, other studies showed that CRH had an antipyretic action when administered centrally into the third cerebral ventricle, which was believed to be due to subsequent release of glucocorticoids£Û4,5£Ý. These data suggested that central CRH might have two independent thermoregulatory actions.

¡¡In this study, intracerebroventricular administration of CRH produced significant increases in body temperature of rats, which is similar to those reported previously£Û2£Ý. Furthermore, central injection of CRH significantly increased the content of AVP in the VSA, microinjection of AVP V1 receptor antagonist into the VSA markedly enhanced the febrile response to central injection of CRH, and injection of AVP into the VSA significantly inhibited fever evoked by central administration of CRH. Although the releaser of AVP responsible for the antipyretic actions of AVP in the VSA has not been well established, the current evidence has shown that AVP acts in the VSA of the brain as a neurotransimitter involved in endogenous antipyresis£Û6£Ý. This, together with the results of the present study, indicates that central CRH stimulates the release of AVP in the VSA during fever, and in turn, endogenous AVP in the VSA has an antipyretic action on the CRHª²induced fever. This may be one of the reasons why the central CRH has an antipyretic action.

In conclusion, CRH may be a dual action mediator in the regulation of body temperature during fever. On the one hand, CRH itself induces an increase in body temperature. On the other hand, CRH stimulates the release of glucocorticoids and AVP to limit the febrile response.

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Received 2001ª²02ª²28Accepted 2001ª²05ª²17

This study was supported by the National Natural Science Foundation of China (No.39700055).

ª³Corresponding author. Tel: 020ª²85220269; Fax: 020ª²85220269; Eª²mail:obszb@jnu.edu.cn