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The effect of a serotonin uptake inhibitor (Lilly 110140) on the secretion of prolactin in the rat
Life Sciences vol . 17, pp . 1141-1144 Printed in the U.S .A .
Pergamon Press
TER Km= OP A SSRO'ZONIN UPTA1E -BITOR (L=Y 110140) On THE SECRETION Or PROLhœIN IN TRE RAT Ladislav MCrulich Department of Physiology, The University of Texas Health Science Center at Dallas, Southwstern Medical School, Dallas, Taxas 75235 (Received is final fore September 8, 1975) SWIARü! Ssrotonin reuptake inhibitor 3-(p-trifloorcoothylphanoxy)-ü-stethyl3-phonylpropylamins (Lilly 110140) is a dose of 10 mV/kg i.p . had no effect an the resting levels of serum . prolactin in normal sale rats . eo~wever, pretreatment of the animals with this drug strongly potentiated the prolactin releasing effect of 5-bydzczytzyvtcphan as wall as the prolactin release induced by ether stress and blood withdrawal . These results indicate that a central serctoninargic mechanism in involved is the stress-induced prolactin release in the rat. . plays as Thori is growing evidence thst a central serotoninergic aechmism important role in the regulation of prolactin secretion. serotonin injected either into the third ventricle (1) or systemically (2) causes as increase of serum pro1actla level.. Systemic injection of a serotonin precursor, 5-hydroxytryptophan (5-WP), has a similar affect (3) . On the contrary, inhibition of serotonin synthesis with g-chlorophonylalani» blocked prolactia release during suckling (4), whereas administra tion of methysergids, a serotonin receptor blocker, inhibited the rip cf prolactin levels following estrogen injection (5) or the stress-induced prolactin release (6) . It was therefore of interest to test the effect of 3-(p-trifluccometbylphenwty)-1i-aethyl-3-phanylpropylamine (Lilly 110140) on the prolactin secretion in resting conditions as well as following stimulation of prolactia releap either by injection of 5-RTP or by stress since this compound has been shown to inhibit specifically serotonin uptake by synaptosomes is the rat brain (7,8) . material and Methods Adult male rata of the Sprague-Davley strain (Simonson Laboratories) weighing around 270 g were used . The animals ware housed is colony cages is groups of 8 is an air-conditioned animal room and they had unlimited access to food and water. The following drugs ware needs 10 mg/kq of Lilly 110140, obtained through the courtesy of Drs. R .W . Fuller and R .S . Bosley of Lilly Research Laboratories ; 5methyiester BC1, 25 and 50 mg/kgt 200 mg/kg of a-methyl-p tyrwinat and 10 mg/kg of mathysergide, kindly provided by Sandox Pharmaowticals . All drugs were dissolved in 0.9% NaCl and injected i.p . is a volume of 0.5 ml . Controls were injected with an equivalent volume of saline . Blood was obtained either from the trunk following decapitation with a 1141
1142
Serotonin Rouptake and Prolactin Secretion
Vol . 17, No . 7
guillotine or from the external jugular win in ether-anesthetized animals . Serum prolactin was determined by the NIAM radioinsounoassay kit with the RP-1 preparation used for standard . All samples were measured in the same assay . The statistical significance of the results was evaluated with the Student's "t" test . The following experimental groups were used : 1) A control group was injected with saline i .p . and killed by decapitation 4 h later. 2) Three groups of 8 esa~i- were injected with Lilly 110140 and killed 2, 4 and 8 h after the injection, respectively . 3) Groups of animals injected with either saline or Lilly 110140 4 h previously received 25 mq/kq or 50 mq/kq of 5-BTP i.p . They were killed 1 h later . 4) One group of animals was injected with saline, another group with 110140 . Four hours after injection the animals of each respective group were bled twice under ether anesthesia within an interval of 10 min. Between sampling the rats were put into individual cages, after the skin incision had been closed with a metal clip .
Lilly
5) a-mothyi-p-tyrosine was injected either into control animals or animals injected with Lilly 110140 4 h proviously . was 6) A group of rats injected with msthysergide 2 h after injection of 110140 and was killed 2 h later. A control group was injected with mothysergide only .
Lilly
Results As can be seen in Table 1, injection of Lilly 110140 had no effect on the nonstress prolactin levels when the values were compared with either the saline injected controls or when the three different times following injection are compared +peon themselves . TABLE 1 Serum prolaçtin ng/ml t SE 17 .8 1 4.13 (8)
Control
Lilly
110140 :
2 hours
13 .9 t 0 .93 (8)
4 hours
9 .6 t 1 .40 (8)
e hours
14 .1 t 2 .42 (8)
Resting serum prolactin levels following injection of Lilly 110140 (10 mg/kg, i.p .) . Animals were killed 2, 4 and 8 hours, respectively, Controls were killed 4 hours followafter the injection of the drug . The numbers in parentheses ing injection of 0.5 ml of saline i .p . indicate the number of animals per group. In control animals, 50 zig/kq of 5-HTP caused a significant rise of prolactin levels whereas 25 mg/kg had no effect (Fig . 1) . In the rats treated with Lilly 110140, already the lower dose of 5-M significantly elevated serum pro lactin levels and the effect of the higher dose was almost tripled with respect to the control group.
Vol . 17, No . 7
Serotonin Reuptake and Prolactin Secretion
1143
FIG. 1 Serum prolactin levels 1 h after injection of 25 and 50 sg/kq of 5-MM in control rats and in rats injected with Lilly 110140 4 h before injection of 5-RM . Eight animals psrgroup were used . * - significant for P<0 .01 against controls .
The animals treated with Lilly 110140 had higher prolactin levels after exposure to ether stress than the corresponding controls (Fig . 2) . In both groups the stress levels ware significantly elevated with respect to the levels of the nonstressed controls . The animal* injected either with saline or with Lilly 110140 4 h before sacrifice were used as controls for the stressed animals. FIG. 2 Effect of ether stress and blood withdrawal on serum prolactin is controls and in animals pretreated with Lilly 110140, respectively . The second sample was taken 10 min after the first sample from tâe sue animals .
Pretreatment of the animals with Lilly 110140 had no effect on the rise of serum prolactin following 01-UT (Table 2) . The same table shows that metbpsergide had no significant effect on serum prolactin either in Lilly 110140treated animals or in controls . TABLE 2 Serumprolactin ng/al t ?E Oontrol
17 .8 1
4 .13 (8)
04er
91 .7 1
8 .59 (8)
9 .6 t
1 .40 (8)
Lilly 110140 Lilly 110140
+ a-M
Nethysergide
Lilly
110140 + Nithysergide
90 .4 1 11 .15 (8) * 11 .3 t
2 .17 (8)
16 .4 3
3 .21 (8)
Seam prolactin levels following injection of a-methyl-p-tyrosine or methysergide either alone or following Lilly 110140 . The animals were killed 1 h following injection of a-NT and 2 h following injection of sothysergide . Lilly 110140 was injected 4 h prior to the injection of either CL-NT or methysergide . * significant for P<0.01 with respect to controls .
1144
Sarotonin Reuptake and Prolactin Secretion
Vol. 17, No . 7
Discussion The results obtained in this short study sight be possibly interpreted in the following manners The resptake of serotonin by the serotoninargic nerve terminals following its release is probably one of the factors which terminate its action at the synapses . It might be therefore expected that an inhibition of this process would enhance the effectiveness of the serotoaiaergic neural mechanisms. The potentiation of the effect of relatively low doses of 5-HTP by Lilly 110140 on seam prolactin obviously fits this concept . The fact that the prolactin releasing effect of stress was also enhanced in this situation, whereas it can be blocked by methysergide (6), lards by inference to the conclusion that a central serotoninergic mechanism is instrumental in the stressinduced release of prolactin. on the other hand, the lack of any affect of either Lilly 110140 or msthyservide or their combinations on the nonstress levels of prolactin probably sagas that the sarotominargic "tonus" affecting the hypothalamic prolactin reg ulating usehana s is -ana-s in resting conditions . The fact that the elevetion of prolactin following removal of the inhibition of prolactin secretion carted by the dopamine-PI: system (10) following administration of O-W was not affected by the serotonin uptake inhibitor also speaks for this asasaption . References 1. 2. 3. 4. 5. 6. 7. 8. 9. 10 .
I.A . EONEtI, R.S . XXCAL, and J.C . PORTM, Endocrinology 88s 1288-1293 (1971) . D.M . LaWSON and R.R . ŒILA, Endocrinology 96s 313-318 (1975) . L.H . LU and J. MEIM , Endoerinolow 93s 152-155 (1973) . C. il Ir OlE, C.A . BIM, J . TElAEEL, and C.H . SAUTER, Neu;oandocrinclogy 13s 213-223 (1M) . L. CALIGRRIS and S . TALEISMIX, J . Zndo=inol . 62s 25-33 (1974) . A. NARCB.3Uß1U1-MW and L . "MILICE, Fed . Proc . 34s 252(abs) (1975) . D.T . VM, J .S . HOM, F .P . BINR9T=, X.L . 1EAD M, and B .B . MOLLOY, Life Sci. 15s 471-479 (1974) . 11 FULLBA, kc .W . PERRY, and B .B . MOLLOY, Life Soi. 15s 1161-1171 (1974) . W. FULLER, B .D . SDWDDY, and 9.3 . MOLLOY, PlOq . of tûe 56th Endocrine Soc. p. A-137 (abs) (1974) . A.O . DQi080, W. BIBHOP, C.P . FAIICLRT, L. i
Pergamon Press
TER Km= OP A SSRO'ZONIN UPTA1E -BITOR (L=Y 110140) On THE SECRETION Or PROLhœIN IN TRE RAT Ladislav MCrulich Department of Physiology, The University of Texas Health Science Center at Dallas, Southwstern Medical School, Dallas, Taxas 75235 (Received is final fore September 8, 1975) SWIARü! Ssrotonin reuptake inhibitor 3-(p-trifloorcoothylphanoxy)-ü-stethyl3-phonylpropylamins (Lilly 110140) is a dose of 10 mV/kg i.p . had no effect an the resting levels of serum . prolactin in normal sale rats . eo~wever, pretreatment of the animals with this drug strongly potentiated the prolactin releasing effect of 5-bydzczytzyvtcphan as wall as the prolactin release induced by ether stress and blood withdrawal . These results indicate that a central serctoninargic mechanism in involved is the stress-induced prolactin release in the rat. . plays as Thori is growing evidence thst a central serotoninergic aechmism important role in the regulation of prolactin secretion. serotonin injected either into the third ventricle (1) or systemically (2) causes as increase of serum pro1actla level.. Systemic injection of a serotonin precursor, 5-hydroxytryptophan (5-WP), has a similar affect (3) . On the contrary, inhibition of serotonin synthesis with g-chlorophonylalani» blocked prolactia release during suckling (4), whereas administra tion of methysergids, a serotonin receptor blocker, inhibited the rip cf prolactin levels following estrogen injection (5) or the stress-induced prolactin release (6) . It was therefore of interest to test the effect of 3-(p-trifluccometbylphenwty)-1i-aethyl-3-phanylpropylamine (Lilly 110140) on the prolactin secretion in resting conditions as well as following stimulation of prolactia releap either by injection of 5-RTP or by stress since this compound has been shown to inhibit specifically serotonin uptake by synaptosomes is the rat brain (7,8) . material and Methods Adult male rata of the Sprague-Davley strain (Simonson Laboratories) weighing around 270 g were used . The animals ware housed is colony cages is groups of 8 is an air-conditioned animal room and they had unlimited access to food and water. The following drugs ware needs 10 mg/kq of Lilly 110140, obtained through the courtesy of Drs. R .W . Fuller and R .S . Bosley of Lilly Research Laboratories ; 5methyiester BC1, 25 and 50 mg/kgt 200 mg/kg of a-methyl-p tyrwinat and 10 mg/kg of mathysergide, kindly provided by Sandox Pharmaowticals . All drugs were dissolved in 0.9% NaCl and injected i.p . is a volume of 0.5 ml . Controls were injected with an equivalent volume of saline . Blood was obtained either from the trunk following decapitation with a 1141
1142
Serotonin Rouptake and Prolactin Secretion
Vol . 17, No . 7
guillotine or from the external jugular win in ether-anesthetized animals . Serum prolactin was determined by the NIAM radioinsounoassay kit with the RP-1 preparation used for standard . All samples were measured in the same assay . The statistical significance of the results was evaluated with the Student's "t" test . The following experimental groups were used : 1) A control group was injected with saline i .p . and killed by decapitation 4 h later. 2) Three groups of 8 esa~i- were injected with Lilly 110140 and killed 2, 4 and 8 h after the injection, respectively . 3) Groups of animals injected with either saline or Lilly 110140 4 h previously received 25 mq/kq or 50 mq/kq of 5-BTP i.p . They were killed 1 h later . 4) One group of animals was injected with saline, another group with 110140 . Four hours after injection the animals of each respective group were bled twice under ether anesthesia within an interval of 10 min. Between sampling the rats were put into individual cages, after the skin incision had been closed with a metal clip .
Lilly
5) a-mothyi-p-tyrosine was injected either into control animals or animals injected with Lilly 110140 4 h proviously . was 6) A group of rats injected with msthysergide 2 h after injection of 110140 and was killed 2 h later. A control group was injected with mothysergide only .
Lilly
Results As can be seen in Table 1, injection of Lilly 110140 had no effect on the nonstress prolactin levels when the values were compared with either the saline injected controls or when the three different times following injection are compared +peon themselves . TABLE 1 Serum prolaçtin ng/ml t SE 17 .8 1 4.13 (8)
Control
Lilly
110140 :
2 hours
13 .9 t 0 .93 (8)
4 hours
9 .6 t 1 .40 (8)
e hours
14 .1 t 2 .42 (8)
Resting serum prolactin levels following injection of Lilly 110140 (10 mg/kg, i.p .) . Animals were killed 2, 4 and 8 hours, respectively, Controls were killed 4 hours followafter the injection of the drug . The numbers in parentheses ing injection of 0.5 ml of saline i .p . indicate the number of animals per group. In control animals, 50 zig/kq of 5-HTP caused a significant rise of prolactin levels whereas 25 mg/kg had no effect (Fig . 1) . In the rats treated with Lilly 110140, already the lower dose of 5-M significantly elevated serum pro lactin levels and the effect of the higher dose was almost tripled with respect to the control group.
Vol . 17, No . 7
Serotonin Reuptake and Prolactin Secretion
1143
FIG. 1 Serum prolactin levels 1 h after injection of 25 and 50 sg/kq of 5-MM in control rats and in rats injected with Lilly 110140 4 h before injection of 5-RM . Eight animals psrgroup were used . * - significant for P<0 .01 against controls .
The animals treated with Lilly 110140 had higher prolactin levels after exposure to ether stress than the corresponding controls (Fig . 2) . In both groups the stress levels ware significantly elevated with respect to the levels of the nonstressed controls . The animal* injected either with saline or with Lilly 110140 4 h before sacrifice were used as controls for the stressed animals. FIG. 2 Effect of ether stress and blood withdrawal on serum prolactin is controls and in animals pretreated with Lilly 110140, respectively . The second sample was taken 10 min after the first sample from tâe sue animals .
Pretreatment of the animals with Lilly 110140 had no effect on the rise of serum prolactin following 01-UT (Table 2) . The same table shows that metbpsergide had no significant effect on serum prolactin either in Lilly 110140treated animals or in controls . TABLE 2 Serumprolactin ng/al t ?E Oontrol
17 .8 1
4 .13 (8)
04er
91 .7 1
8 .59 (8)
9 .6 t
1 .40 (8)
Lilly 110140 Lilly 110140
+ a-M
Nethysergide
Lilly
110140 + Nithysergide
90 .4 1 11 .15 (8) * 11 .3 t
2 .17 (8)
16 .4 3
3 .21 (8)
Seam prolactin levels following injection of a-methyl-p-tyrosine or methysergide either alone or following Lilly 110140 . The animals were killed 1 h following injection of a-NT and 2 h following injection of sothysergide . Lilly 110140 was injected 4 h prior to the injection of either CL-NT or methysergide . * significant for P<0.01 with respect to controls .
1144
Sarotonin Reuptake and Prolactin Secretion
Vol. 17, No . 7
Discussion The results obtained in this short study sight be possibly interpreted in the following manners The resptake of serotonin by the serotoninargic nerve terminals following its release is probably one of the factors which terminate its action at the synapses . It might be therefore expected that an inhibition of this process would enhance the effectiveness of the serotoaiaergic neural mechanisms. The potentiation of the effect of relatively low doses of 5-HTP by Lilly 110140 on seam prolactin obviously fits this concept . The fact that the prolactin releasing effect of stress was also enhanced in this situation, whereas it can be blocked by methysergide (6), lards by inference to the conclusion that a central serotoninergic mechanism is instrumental in the stressinduced release of prolactin. on the other hand, the lack of any affect of either Lilly 110140 or msthyservide or their combinations on the nonstress levels of prolactin probably sagas that the sarotominargic "tonus" affecting the hypothalamic prolactin reg ulating usehana s is -ana-s in resting conditions . The fact that the elevetion of prolactin following removal of the inhibition of prolactin secretion carted by the dopamine-PI: system (10) following administration of O-W was not affected by the serotonin uptake inhibitor also speaks for this asasaption . References 1. 2. 3. 4. 5. 6. 7. 8. 9. 10 .
I.A . EONEtI, R.S . XXCAL, and J.C . PORTM, Endocrinology 88s 1288-1293 (1971) . D.M . LaWSON and R.R . ŒILA, Endocrinology 96s 313-318 (1975) . L.H . LU and J. MEIM , Endoerinolow 93s 152-155 (1973) . C. il Ir OlE, C.A . BIM, J . TElAEEL, and C.H . SAUTER, Neu;oandocrinclogy 13s 213-223 (1M) . L. CALIGRRIS and S . TALEISMIX, J . Zndo=inol . 62s 25-33 (1974) . A. NARCB.3Uß1U1-MW and L . "MILICE, Fed . Proc . 34s 252(abs) (1975) . D.T . VM, J .S . HOM, F .P . BINR9T=, X.L . 1EAD M, and B .B . MOLLOY, Life Sci. 15s 471-479 (1974) . 11 FULLBA, kc .W . PERRY, and B .B . MOLLOY, Life Soi. 15s 1161-1171 (1974) . W. FULLER, B .D . SDWDDY, and 9.3 . MOLLOY, PlOq . of tûe 56th Endocrine Soc. p. A-137 (abs) (1974) . A.O . DQi080, W. BIBHOP, C.P . FAIICLRT, L. i