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Separation of pressor and antidiuretic effects of intraventricular bradykinin
SEPARATION OF PRESSOR AND ANTIDIURETIC EFFECTS OF INTRAVENTRICULAR BRADYKININ W. E. HOFFMAN and P. G. SCHMID Department of Medical Physiology. Texas A & M University College of Medicine. College Station. TX 77843. U.S.A. and Cardiovascular Division. Department of Internal Medicine. University of Iowa. Iowa City. iA 5224O.‘U.S.A. (Accrpred
30 January
1978)
Summary-Intraventricular (i.v.t.) injections of bradykinin have been reported to produce centrally mediated pressor effects. Here it is reported that intraventricular bradykinin infusions produce antidiuretic effects in addition to pressor effects previously reported. Median eminence lesions block the antidiuretic but not the pressor effects of central bradykinin, suggesting that ADH release is the major component of antidiuresis but that other central mechanisms, probably sympathetic outflow, mediate the blood pressure increase. The similarity of central effects seen with bradykinin and angiotensin II suggests a common mechanism of action within the central nervous system.
It has been reported by Lambert and Lang (1970) and Correa and Gtaeff (1974) that intraventricular (i.v.t.) injections of bradykinin will produce pressor effects in the rat. These effects are similar to those of carbachol and angiotensin II, which also produce centrally mediated pressor effects in the unanaesthetized rat (Hoffman. Phillips. Schmid, Falcon and Weet, 1977: Severs, Summy-Long, Daniels-Severs and Connor, 1971). In addition to producing pressor effects, the latter two drugs have also been shown to produce dose-dependent antidiuretic effects which suggest antidiuretic hormone (ADH) release (Kuhn, 1974; Hoffman er al., 1977). It was the purpose of these experiments to determine whether intraventricular bradykinin produces antidiuretic effects similar to those seen with carbachol and angiotensin II or if pressor responses to central bradykinin occur independently of ADH release. METHODS
Male Sprague-Dawley rats were implanted with chronic 23-gauge third ventricular cannulae 3 days before testing under chloral hydrate anaesthesia. On the day of testing each rat was anaesthetized with halothane and implanted with PESO catheters in the femoral artery and vein. These catheters were fed subcutaneously through a small incision in the back. The urinary bladder was also catheterized with PE90 tubing via an abdominal approach. Following this surgery the rats were allowed to recover from the halothane anaesthesia. An intravenous (iv.) infusion (0.20 ml/min) of hydrating solution containing 10 mM sodium chforide, 5mM sodium bicarbonate and 135mM glucose was started and continued for the duration of the experiment. During the experiment, blood pressure, heart rate, urine flow and urine conductance were continuously recorded on a Beckman Dynograph recorder. The rats were confined to a X.P.17’12~-*
25 x 10 x 9cm black plastic box but were not restrained. Bradykinin was obtained from Sigma and was dissolved in artificial cerebrospinal fluid (CSF, Travenol labs). Intraventricular injections of bradykinin were given in 5 ~1. volumes through a 30-gauge injection cannula designed to fit inside and to end flush with the chronically implanted third cerebral ventricular cannulae. A 25~1 Hamilton syringe, used for microjnjections, was connected to the injection cannula with PElO tubing. Vehicle control intraventricular injections were given in the same volume as in bradykinin tests (5~1). In order to compare antidiuretic responses to intraventricular bradykinin with antidiuretic hormone (ADH) standards, ADH was added to the hydrating infusion in doses of 0.1, 0.5, 2.0 and 4.0mU at a rate of 0.1 mUisec. Two to four doses of ADH were tested in each rat. Saline vehicles were used as a control for volume. Antidiuretic responses are defined here as a simultaneous decrease in urine flow and an increase in urine conductance (electrolyte concentration). For analysis, changes in urine conductance are reported for antidiuretic responses, a method used in rat ADH bioassays (Pliska and Rychlik, 1967). Median eminence lesions were performed with an insulated stainless-steel electrode using a 2 mA. 1S-set anodal current. The ear bar of the stereostaxic instrument served as the indifferent electrode. These lesions were carried out under choral hydrate anaesthesia at the time of the third ventricular cannula implantation. Protocol for testing median eminence-lesioned rats was the same as in control animals. Following testing, the brains of these rats were examined histologically to verify the extent of the lesion. An adequate lesion was assessed as SCrlOO% destruction of the median eminence. Sham-lesioned rats received the same surgical procedure but were not lesioned. Results are reported as means & SE. and comparisons were made with a Student’s t-test.
HOFFMAN
W. E.
and P. G.
SCHMID
200
Blood pressure
I00
F
(mmHg)
5~
BK
0 E
E--r-----4:0
400
Heart rate (rnln-‘)
360
240
R_.1lIIIIIIIIIllIllllllllllllllllllllllllllllHlllsl
Urine flow (drops1 Urbne conductance
z”&?$?$$-
(PMhos)
IO
5
0
Time
15
20
(minutes)
Fig. 1. Cardiovascular and antidiuretic effect of 5 pg bradykinin (BK) intraventricular shown are urine conductance curves. measured in PMhos, produced by intravenous 2 and 4mU ADH in the same rat.
injection. Also infusions of 0.5.
L!dd NORMAL
(n=21)
300
r
9,
1 5, z! r” ZE Pss
a?
0
CSF
0.05
10
Bradykmm
50
200 -
*
$$
a-100._ .s E q
mf =2 .s - -30 t: : Jz
10
0
E:
$2
z 57
**
2 0
30
-60
* CSF
0.05
10
5 0
Bradykinm
(pgl
MEDIAN
0
(pg)
EMINENCE
IY CSF
= 0.05
1.0
5 0
Bradykmm
(pg)
* *tt
01
1 520
mU ADH
( n= 5 )
LESIONS
*t
50 iJg Eradykmm
0 010
pz.05 pe.01 PC
,001
*** I
sow2 Sradykmm
0 010520 mU ADH
Fig. 2. Blood pressure, heart rate and antidiuretic effects of bradykinin intraventricular infusions in normal and median eminence-lesioned rats. Also shown are antidiuretic responses to intravenous ADH infusions in both groups of rats. Conductance values are reported in PMhos. Values reported as mean f S.E. and comparisons made with Student’s r-test. Significant levels of responses in normal rats indicate difference from cerebrospinal fluid control infusions. Significant levels of median eminence lesion data indicate difference from same dose given in normal rats. All rats received both intraventricular bradykinin and intravenous ADH infusions.
Bradykinin intraventricular RESULTS There was no difference in pressor or antidiuretic responses between 7 sham-median eminence lesioned rats and 14 other control animals. Therefore, both groups were combined for data analysis. Mean blood pressure and heart rate of the 21 rats tested was 111 +_ 3 mmHg and 379 + 9 min-‘. In these rats the continuous hydrating infusion produced a diuresis which was low in conductance. Intravenous infusions of ADH produced decreases in urine flow and dosedependent increases in urine conductance (Figs 1 and 2). Pressor effects of intravenous ADH infusions were: 0.1mU=O~OmmHg(n=8),0.5mU=2~lmmHg (n=15),2mU=7+2mmHg(n=21),4mU=12f2 mmHg (n = 12). Intraventricular bradykinin infusions produced dose-dependent increases in blood pressure and antidiuresis (Figs 1 and 2). In several rats an initial, transient tachycardia was observed although this changed to a longer-term bradycardia for the highest dose of bradykinin. Intravenous infusions of 5 pg bradykinin (n = 8) produced a decrease in blood pressure (-9 +_ 3 mmHg), tachycardia (67 & 16 mini) and a decrease rather than an increase in urine conductance (- 30 +_ 10 Mhos). In 5 rats with median eminence lesions mean blood pressure and heart rate was 110 + 7mmHg and 427 f 33 min-‘. not significantly different from normal rats (P > 0.05). Intravenous ADH infusions produced dose-dependent antidiuretic effects in these rats which were not significantly different from control animals (Fig. 2). Intraventricular infusions of 5 pg bradykinin in these rats produced increases in blood pressure and bradycardia as in normal rats. Pressor responses to intraventricular bradykinin infusions were reduced after median eminence lesions but this reduction was not significant compared to nonlesioned rats. Antidiuretic responses to central bradykinin infusions were abolished in median eminence lesioned rats. DISCUSSION These results indicate that intraventricular bradykinin produces centrally mediated antidiuretic effects in addition to pressor responses. Intravenous infusions of bradykinin produce neither of these responses. The antidiuretic effects are probably due to ADH release since they are abolished by median eminence lesions, while the pressor effects are not significantly changed by this treatment. This suggests that central bradykinin can initiate ADH release as well as stimulating central sympathetic outflow but the pathways for these responses are separate. The pressor effects seen here to intraventricular injections of bradykinin are in agreement with the previous report by Lambert and Lang (1970) and Correa and Graeff (1974). The antidiuretic effects of intraventricular bradykinin have not been reported before. Since the components of the kinin system have been identified in brain tissue by Hori (1968) and Car-
1001
margo and Graeff (1969) these responses may represent physiological mechanisms of the central kinin system. The doses used here and in previous studies are large, however, and it is not known if these pressor and antidiuretic responses represent physiological or pharmacological effects. Another peptide with similar central actions to bradykinin is angiotensin II. Angiotensin II will also produce dose-dependent pressor and antidiuretic effects in unanaesthetized rats although lower doses of angiotensin are required to produce similar responses (Hoffman et al., 1977). In addition to producing pressor and antidiuretic effects, the pressor responses to central bradykinin and to angiotensin II are both blocked by central pretreatment with phentolamine, an sc-adrenergic blocker (Correa and Graeff, 1974; Severs, Summy-Long, Daniels-Severs and Connor, 1970). This suggests that central noradrenergic mechanisms in some way modulate both bradykinin and angiotensin II pressor effects. The relationship between central sympathetic stimulation and ADH release is unclear. At present it appears that drugs which stimulate one response in the brain will also initiate the second. These effects can be separated, for example, by median eminence lesions. Median eminence lesions were used in these studies to block ADH release. The adequacy of these lesions was supported by histological examination. Antidiuretic effects were inhibited to intraventricular bradykinin in lesioned animals compared to control rats while pressor effects were not significantly changed. Release of ADH may have played a small role in the pressor effects of intraventricular bradykinin as evidenced by the blood pressure responses to intravenous ADH infusions. However, the evidence suggests that sympathetic mechanisms play the major role in the blood pressure increase to intraventricular bradykinin. The specificity of the effects of median eminence lesions is supported by the fact that pressor responses to intraventricular bradykinin remain intact while antidiuresis is abolished. Correa and Graeff (1975) have shown the importance of the lateral septal region in pressor effects to central bradykinin when they demonstrated that lateral septal lesions abolish pressor effects to intraventricular bradykinin. This suggests that the lateral septal region mediates sympathetic outflow to central bradykinin stimulation. The importance of the lateral septal region in mediating antidiuretic effects of intraventricular bradykinin is not known. In conclusion, these results demonstrate that intraventricular bradykinin infusions produce antidiuretic as well as pressor effects in the unanaesthetized rat. The fact that antidiuretic effects of central bradykinin infusions are abolished by median eminence lesions while pressor effects are not significantly changed suggests that ADH release mediates the antidiuresis. Pressor effects are most likely due to sympathetic outflow mediated by a-adrenergic mechanisms and receptor sites in the lateral septal region.
1002
W. E. HOFFMAN and P. G. SCHMID
AcktloMledyemenrs--This work was supported by Program Project Grant No. HL 14388 to PG and NIH Research Fellowship No. HLOSS3S to WEH.
REFERENCES Carmargo, A. C. M. and Graeff, F. G. (1969). Subcellular distribution and properties of the bradykinin in activation system in rabbit brain homogenates. Biochrm. Pharmac. 18: 548-549. Correa. F. M. A. and Graeff. F. G. (1974). Central mechanisms of the hypertensive action of intraventricular bradykinin in the unanaesthetized rat. .Ric,uropharmacoltg~ 13: 65-75.
Correa, F. M. A. and Graeff, F. Cr. (1975). Central site of the hypertensive action of bradykinin. J. Pharmac. PXD. Thu.
192: 67@676.
Hoffman. W. E., Phillips. M. I., Schmid. P. G.. Falcon, J. and Weet. J. F. (1977). Antidiuretic hormone release
and the pressor response to central angiotensin II and cholinergic stimulation, Nuuropharmacology 16: 463-472. Hori. S. (1968). The presence of bradykinin like polypeptides. kinin releasing and destroying activity in brain. Jap. J. Physiol. 18: 772-787. Kuhn, E. R. (1974). Cholinergic and adrenergic release mechanism for vasopressin in the male rat: a study with injections of neurotransmitters and blocking agents into the third ventricle. Neuroendocrinology 16: 2555264. Lambert. G. A. and Lang, W. J. (1970). The effects of bradykinin and eledosin injected into the cerebral ventricles of conscious rats. Eur. J. Pharmac. 9: 383-386. Pliska. V. and Rychlik, 1. (1967). Determination of antidiuretic activity in the rat for structural analogues of the neurohypophysical hormones. Acra endocr. 54: I29m 140. Severs. W. B., Summy-Long. J.. Daniels-Severs. A. and Connor. J. D. (1971). Influence of adrenergic blocking drugs on central angiotensin effects. Pharmacology 5: 205m 2 14.
30 January
1978)
Summary-Intraventricular (i.v.t.) injections of bradykinin have been reported to produce centrally mediated pressor effects. Here it is reported that intraventricular bradykinin infusions produce antidiuretic effects in addition to pressor effects previously reported. Median eminence lesions block the antidiuretic but not the pressor effects of central bradykinin, suggesting that ADH release is the major component of antidiuresis but that other central mechanisms, probably sympathetic outflow, mediate the blood pressure increase. The similarity of central effects seen with bradykinin and angiotensin II suggests a common mechanism of action within the central nervous system.
It has been reported by Lambert and Lang (1970) and Correa and Gtaeff (1974) that intraventricular (i.v.t.) injections of bradykinin will produce pressor effects in the rat. These effects are similar to those of carbachol and angiotensin II, which also produce centrally mediated pressor effects in the unanaesthetized rat (Hoffman. Phillips. Schmid, Falcon and Weet, 1977: Severs, Summy-Long, Daniels-Severs and Connor, 1971). In addition to producing pressor effects, the latter two drugs have also been shown to produce dose-dependent antidiuretic effects which suggest antidiuretic hormone (ADH) release (Kuhn, 1974; Hoffman er al., 1977). It was the purpose of these experiments to determine whether intraventricular bradykinin produces antidiuretic effects similar to those seen with carbachol and angiotensin II or if pressor responses to central bradykinin occur independently of ADH release. METHODS
Male Sprague-Dawley rats were implanted with chronic 23-gauge third ventricular cannulae 3 days before testing under chloral hydrate anaesthesia. On the day of testing each rat was anaesthetized with halothane and implanted with PESO catheters in the femoral artery and vein. These catheters were fed subcutaneously through a small incision in the back. The urinary bladder was also catheterized with PE90 tubing via an abdominal approach. Following this surgery the rats were allowed to recover from the halothane anaesthesia. An intravenous (iv.) infusion (0.20 ml/min) of hydrating solution containing 10 mM sodium chforide, 5mM sodium bicarbonate and 135mM glucose was started and continued for the duration of the experiment. During the experiment, blood pressure, heart rate, urine flow and urine conductance were continuously recorded on a Beckman Dynograph recorder. The rats were confined to a X.P.17’12~-*
25 x 10 x 9cm black plastic box but were not restrained. Bradykinin was obtained from Sigma and was dissolved in artificial cerebrospinal fluid (CSF, Travenol labs). Intraventricular injections of bradykinin were given in 5 ~1. volumes through a 30-gauge injection cannula designed to fit inside and to end flush with the chronically implanted third cerebral ventricular cannulae. A 25~1 Hamilton syringe, used for microjnjections, was connected to the injection cannula with PElO tubing. Vehicle control intraventricular injections were given in the same volume as in bradykinin tests (5~1). In order to compare antidiuretic responses to intraventricular bradykinin with antidiuretic hormone (ADH) standards, ADH was added to the hydrating infusion in doses of 0.1, 0.5, 2.0 and 4.0mU at a rate of 0.1 mUisec. Two to four doses of ADH were tested in each rat. Saline vehicles were used as a control for volume. Antidiuretic responses are defined here as a simultaneous decrease in urine flow and an increase in urine conductance (electrolyte concentration). For analysis, changes in urine conductance are reported for antidiuretic responses, a method used in rat ADH bioassays (Pliska and Rychlik, 1967). Median eminence lesions were performed with an insulated stainless-steel electrode using a 2 mA. 1S-set anodal current. The ear bar of the stereostaxic instrument served as the indifferent electrode. These lesions were carried out under choral hydrate anaesthesia at the time of the third ventricular cannula implantation. Protocol for testing median eminence-lesioned rats was the same as in control animals. Following testing, the brains of these rats were examined histologically to verify the extent of the lesion. An adequate lesion was assessed as SCrlOO% destruction of the median eminence. Sham-lesioned rats received the same surgical procedure but were not lesioned. Results are reported as means & SE. and comparisons were made with a Student’s t-test.
HOFFMAN
W. E.
and P. G.
SCHMID
200
Blood pressure
I00
F
(mmHg)
5~
BK
0 E
E--r-----4:0
400
Heart rate (rnln-‘)
360
240
R_.1lIIIIIIIIIllIllllllllllllllllllllllllllllHlllsl
Urine flow (drops1 Urbne conductance
z”&?$?$$-
(PMhos)
IO
5
0
Time
15
20
(minutes)
Fig. 1. Cardiovascular and antidiuretic effect of 5 pg bradykinin (BK) intraventricular shown are urine conductance curves. measured in PMhos, produced by intravenous 2 and 4mU ADH in the same rat.
injection. Also infusions of 0.5.
L!dd NORMAL
(n=21)
300
r
9,
1 5, z! r” ZE Pss
a?
0
CSF
0.05
10
Bradykmm
50
200 -
*
$$
a-100._ .s E q
mf =2 .s - -30 t: : Jz
10
0
E:
$2
z 57
**
2 0
30
-60
* CSF
0.05
10
5 0
Bradykinm
(pgl
MEDIAN
0
(pg)
EMINENCE
IY CSF
= 0.05
1.0
5 0
Bradykmm
(pg)
* *tt
01
1 520
mU ADH
( n= 5 )
LESIONS
*t
50 iJg Eradykmm
0 010
pz.05 pe.01 PC
,001
*** I
sow2 Sradykmm
0 010520 mU ADH
Fig. 2. Blood pressure, heart rate and antidiuretic effects of bradykinin intraventricular infusions in normal and median eminence-lesioned rats. Also shown are antidiuretic responses to intravenous ADH infusions in both groups of rats. Conductance values are reported in PMhos. Values reported as mean f S.E. and comparisons made with Student’s r-test. Significant levels of responses in normal rats indicate difference from cerebrospinal fluid control infusions. Significant levels of median eminence lesion data indicate difference from same dose given in normal rats. All rats received both intraventricular bradykinin and intravenous ADH infusions.
Bradykinin intraventricular RESULTS There was no difference in pressor or antidiuretic responses between 7 sham-median eminence lesioned rats and 14 other control animals. Therefore, both groups were combined for data analysis. Mean blood pressure and heart rate of the 21 rats tested was 111 +_ 3 mmHg and 379 + 9 min-‘. In these rats the continuous hydrating infusion produced a diuresis which was low in conductance. Intravenous infusions of ADH produced decreases in urine flow and dosedependent increases in urine conductance (Figs 1 and 2). Pressor effects of intravenous ADH infusions were: 0.1mU=O~OmmHg(n=8),0.5mU=2~lmmHg (n=15),2mU=7+2mmHg(n=21),4mU=12f2 mmHg (n = 12). Intraventricular bradykinin infusions produced dose-dependent increases in blood pressure and antidiuresis (Figs 1 and 2). In several rats an initial, transient tachycardia was observed although this changed to a longer-term bradycardia for the highest dose of bradykinin. Intravenous infusions of 5 pg bradykinin (n = 8) produced a decrease in blood pressure (-9 +_ 3 mmHg), tachycardia (67 & 16 mini) and a decrease rather than an increase in urine conductance (- 30 +_ 10 Mhos). In 5 rats with median eminence lesions mean blood pressure and heart rate was 110 + 7mmHg and 427 f 33 min-‘. not significantly different from normal rats (P > 0.05). Intravenous ADH infusions produced dose-dependent antidiuretic effects in these rats which were not significantly different from control animals (Fig. 2). Intraventricular infusions of 5 pg bradykinin in these rats produced increases in blood pressure and bradycardia as in normal rats. Pressor responses to intraventricular bradykinin infusions were reduced after median eminence lesions but this reduction was not significant compared to nonlesioned rats. Antidiuretic responses to central bradykinin infusions were abolished in median eminence lesioned rats. DISCUSSION These results indicate that intraventricular bradykinin produces centrally mediated antidiuretic effects in addition to pressor responses. Intravenous infusions of bradykinin produce neither of these responses. The antidiuretic effects are probably due to ADH release since they are abolished by median eminence lesions, while the pressor effects are not significantly changed by this treatment. This suggests that central bradykinin can initiate ADH release as well as stimulating central sympathetic outflow but the pathways for these responses are separate. The pressor effects seen here to intraventricular injections of bradykinin are in agreement with the previous report by Lambert and Lang (1970) and Correa and Graeff (1974). The antidiuretic effects of intraventricular bradykinin have not been reported before. Since the components of the kinin system have been identified in brain tissue by Hori (1968) and Car-
1001
margo and Graeff (1969) these responses may represent physiological mechanisms of the central kinin system. The doses used here and in previous studies are large, however, and it is not known if these pressor and antidiuretic responses represent physiological or pharmacological effects. Another peptide with similar central actions to bradykinin is angiotensin II. Angiotensin II will also produce dose-dependent pressor and antidiuretic effects in unanaesthetized rats although lower doses of angiotensin are required to produce similar responses (Hoffman et al., 1977). In addition to producing pressor and antidiuretic effects, the pressor responses to central bradykinin and to angiotensin II are both blocked by central pretreatment with phentolamine, an sc-adrenergic blocker (Correa and Graeff, 1974; Severs, Summy-Long, Daniels-Severs and Connor, 1970). This suggests that central noradrenergic mechanisms in some way modulate both bradykinin and angiotensin II pressor effects. The relationship between central sympathetic stimulation and ADH release is unclear. At present it appears that drugs which stimulate one response in the brain will also initiate the second. These effects can be separated, for example, by median eminence lesions. Median eminence lesions were used in these studies to block ADH release. The adequacy of these lesions was supported by histological examination. Antidiuretic effects were inhibited to intraventricular bradykinin in lesioned animals compared to control rats while pressor effects were not significantly changed. Release of ADH may have played a small role in the pressor effects of intraventricular bradykinin as evidenced by the blood pressure responses to intravenous ADH infusions. However, the evidence suggests that sympathetic mechanisms play the major role in the blood pressure increase to intraventricular bradykinin. The specificity of the effects of median eminence lesions is supported by the fact that pressor responses to intraventricular bradykinin remain intact while antidiuresis is abolished. Correa and Graeff (1975) have shown the importance of the lateral septal region in pressor effects to central bradykinin when they demonstrated that lateral septal lesions abolish pressor effects to intraventricular bradykinin. This suggests that the lateral septal region mediates sympathetic outflow to central bradykinin stimulation. The importance of the lateral septal region in mediating antidiuretic effects of intraventricular bradykinin is not known. In conclusion, these results demonstrate that intraventricular bradykinin infusions produce antidiuretic as well as pressor effects in the unanaesthetized rat. The fact that antidiuretic effects of central bradykinin infusions are abolished by median eminence lesions while pressor effects are not significantly changed suggests that ADH release mediates the antidiuresis. Pressor effects are most likely due to sympathetic outflow mediated by a-adrenergic mechanisms and receptor sites in the lateral septal region.
1002
W. E. HOFFMAN and P. G. SCHMID
AcktloMledyemenrs--This work was supported by Program Project Grant No. HL 14388 to PG and NIH Research Fellowship No. HLOSS3S to WEH.
REFERENCES Carmargo, A. C. M. and Graeff, F. G. (1969). Subcellular distribution and properties of the bradykinin in activation system in rabbit brain homogenates. Biochrm. Pharmac. 18: 548-549. Correa. F. M. A. and Graeff. F. G. (1974). Central mechanisms of the hypertensive action of intraventricular bradykinin in the unanaesthetized rat. .Ric,uropharmacoltg~ 13: 65-75.
Correa, F. M. A. and Graeff, F. Cr. (1975). Central site of the hypertensive action of bradykinin. J. Pharmac. PXD. Thu.
192: 67@676.
Hoffman. W. E., Phillips. M. I., Schmid. P. G.. Falcon, J. and Weet. J. F. (1977). Antidiuretic hormone release
and the pressor response to central angiotensin II and cholinergic stimulation, Nuuropharmacology 16: 463-472. Hori. S. (1968). The presence of bradykinin like polypeptides. kinin releasing and destroying activity in brain. Jap. J. Physiol. 18: 772-787. Kuhn, E. R. (1974). Cholinergic and adrenergic release mechanism for vasopressin in the male rat: a study with injections of neurotransmitters and blocking agents into the third ventricle. Neuroendocrinology 16: 2555264. Lambert. G. A. and Lang, W. J. (1970). The effects of bradykinin and eledosin injected into the cerebral ventricles of conscious rats. Eur. J. Pharmac. 9: 383-386. Pliska. V. and Rychlik, 1. (1967). Determination of antidiuretic activity in the rat for structural analogues of the neurohypophysical hormones. Acra endocr. 54: I29m 140. Severs. W. B., Summy-Long. J.. Daniels-Severs. A. and Connor. J. D. (1971). Influence of adrenergic blocking drugs on central angiotensin effects. Pharmacology 5: 205m 2 14.