Endocrine, vol. 7, no. 2, 177-182, October 1997 0969-711X/97/7:177-182/$9.50 9 1997 by Humana PressInc. All rights of any nature whatsoever reserved. Dopamine D 2 Receptor Mediates Both Inhibitory and Stimulatory Actions on Prolactin Release A. Chang/S. H. Shin/and S. C. Pang 2 Department of IPhysiology, and 2Anatomy and Cell Biology, Queen's University, Kingston, Ontario, Canada Dopamine is considered to be the major physiological tonic inhibitor of prolactin release, yet there is increas- ing evidence showing that it can also stimulate prolac- tin release from lactotrophs. In primary cultured lactotrophs, the major dopamine receptors responsible for inhibiting prolactin release are dopamine D 2 recep- tors. A dopamine receptor subtype may be responsible for the stimulatory action, yet one cannot exclude the possibility that a dopamine D 2 receptor can play dual roles. This study was therefore undertaken to investi- gate if dopamine both stimulates and inhibits prolactin secretion through activation of the same dopamine D 2 receptor. GH4ZR 7 cells, which have only one type of dopamine receptors--D2s , were perifused with dif- ferent concentrations of dopamine, and the perifusate was assayed for prolactin; 10 -7 mol/L dopamine stimu- lated prolactin release (p < 0.05; n -- 5), whereas 5 • 10 -4 mol/L dopamine inhibited prolactin secretion (p< 0.05; n -- 5). In the pertussis toxin-treated cells, 10 -7 mol/L dopamine stimulated prolactin release (p < 0.05; n = 5), and 5 • 10 -4 mol/I, dopamine did not significantly change the rate of prolactin release. These results indi- cate that both the stimulatory and inhibitory actions of dopamine are likely mediated by the same D 2 receptor subtype, since GH4ZR 7 cells express only D2s recep- tors. They also confirm that the inhibitory action of dopamine is mediated through a G i protein; and the stimulatory action of dopamine is mediated through a PTX-insensitive pathway. These findings suggest that D 2 receptors are coupled to both G i and G s proteins. Key Words: Dopamine; D2receptor; prolactin; G H4ZR7; pertussis toxin. Introduction Prolactin is an important hormone involved in the regu- lation of a diversity of physiological processes, including Received April 21, 1997; Revised May 23, 19997; Accepted May 23, 1997. Author to whom all correspondence and reprint requests should be addressed: Dr. S. H. Shin, Department of Physiology, Queen's University, Botterell Hall, Kingston, Ontario, Canada K7L 3N6. E-mail address: shins@post. queensu.ca milk secretion, reproduction, immune responses, osmo- regulation, and growth promotion (I). It is mainly secreted from lactotrophs in the anterior pituitary gland (2), and its release is predominantly under inhibitory control, whereas occasional bursts of prolactin secretion, such as during stress, are likely induced by putative prolactin-releasing factors (3). Dopamine is the major tonic inhibitory regula- tor of prolactin synthesis and release. It has been consid- ered the physiological prolactin release-inhibiting factor (PIF) (4), yet some studies showed that dopamine can also stimulate prolactin release fi'om lactotrophs (1,5-9). Although the majority of dopamine D 2 receptors in primary cultured lactotrophs are inhibitory (7,10-12), there are one or more D 2 receptor subtypes that act as mediators in the stimulatory action of dopamine (7,13). However, one can- not exclude the possibility that a dopamine D 2 receptor can play dual (stimulatory and inhibitory) roles. We therefore have chosen for this study the GH4ZR 7 cell line, which, among all dopamine receptor subtypes, has only the trans- fected D~ receptors (Dzs) (11) to find out if activation of dopamine D 2 receptor only inhibits prolactin release or both stimulates and inhibits prolactin secretion. Results Thyrotropin releasing hormone (TRH) was used in the study as a reference control to ensure that the cells were responding properly to secretagog. In the GH4ZR7 cells, perifusion of TRH (10-6 tool/L) caused a significant increase of prolactin release, in both the control and the pertussis toxin-(PTX) treated cells as expected (Fig. 1A,B). The peak prolactin concentration induced by TRH in the nontreated group (51.7 + 3.3 ng/mL; 268 + 30% of basal level) (mean _+ SEM) (Fig. I A) was not significantly different from that of the PTX-treated group (49.1 _+ 3.4 ng/mL; 263 + 28% of basal level) (Fig. 1B), indicating that PTX had no effect on the stimulatory action of TRH on prolactin release. When GH4ZR 7 cells were treated with 10 -7 mol/L dopamine, the prolactin release was increased from a basal concentration of 23.3 _+ 2.9 ng/mL (100%) (area under the curve [AUC] -- 72.7 _+ 9.1) to a peak concentration of 31.0 +1.6 ng/mL or 125 + 10% of basal concentration (AUC = 102.6 + 6.9) (mean + SEM) (p < 0.05; n = 5) and then returned to a baseline concentration during dopamine 177