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Endocrinology Vol. 143, No. 3 744-746
Copyright © 2002 by The Endocrine Society

INTRACELLULAR SIGNAL SYSTEMS

Editorial: Dopamine-D2-Mediated Inhibition of TRH-Induced PLC Activation in Pituitary Cells—Direct or Indirect?

Paul R. Albert

Ottawa Health Research Institute (Neuroscience), University of Ottawa, Ottawa, Ontario K1H-8M5, Canada

Address all correspondence and requests for reprints to: Paul R. Albert, Ottawa Health Research Institute (Neuroscience), 451 Smyth Road, Ottawa, Ontario K1H-8M5, Canada. E-mail: . palbert{at}uottawa.ca


   Introduction
Top
 Introduction
References
 
The dopamine-D2 receptor is the primary inhibitory regulator of PRL synthesis and secretion from pituitary lactotrophs in vivo (1, 2). Recently, gene knockout studies in mice have illustrated the role of dopamine-D2 receptors in regulation of PRL secretion and synthesis as well as pituitary development (3, 4, 5). Mice deficient in D2 receptors were hyperprolactinemic, with reduced gonad size in both sexes and females that develop uterine adenomyosis, probably resulting from chronic elevation of PRL. In mice deficient in dopamine-D2 receptors, there was pituitary hyperplasia due to proliferation of lactotrophs leading to pituitary adenomas (3, 4, 5). Conversely, augmentation of dopamine release in mice lacking the dopamine transporter, which mediates reuptake of dopamine and termination of its actions, produced an opposite pituitary phenotype (6). These mice displayed hypotrophic pituitaries due to a lack of somatotrophs and lactotrophs and had a reduction secretogogue-induced but not basal PRL and GH secretion, leading to dwarfism. These examples illustrate the important role of D2 receptors in negative regulation of lactotroph and somatotroph development and growth, and in regulating PRL synthesis and secretion in vivo.

Several studies have addressed the mechanisms by which dopamine regulates lactotroph secretion and growth in primary pituitary cell culture or in pituitary cell strains such as GH3 or GH4C1 cells (1, 7). Activation of dopamine-D2 receptors in pituitary cells mediates inhibitory regulation of a variety of signaling pathways. Using distinct Gi proteins, dopamine-D2 receptor activation inhibits basal, forskolin-, or VIP-stimulated cAMP formation in lactotrophs (8, 9, 10). In addition, the D2 receptor opens potassium channels and closes calcium channels using Gi3 and Go, respectively (9, 10, 11, 12). These pathways are blocked by pertussis toxin, a selective inhibitor of Gi/Go proteins. These diverse pathways couple to inhibition of PRL secretion, but the exact roles of these pathways are difficult to define. To inhibit VIP-stimulated PRL secretion, the most likely pathway is mediated by G{alpha}i (G{alpha}i1–3), which couples to inhibition of VIP-induced cAMP formation (8, 9, 10). To inhibit secretion stimulated by BayK8466, a direct L-type calcium channel agonist, the most likely scenario is that dopamine acts by Go-mediated inhibition of calcium channel activation (9, 10, 11, 12). A similar argument can be made for stimulation by high potassium, which is also blocked by L-type calcium channel antagonists. But the mechanism by which dopamine regulates the actions of the physiological secretagogues, such as TRH or angiotensin II, is more complicated.

TRH receptors couple to Gq/11 to stimulate PLC-ß activity, increasing inositol phosphate and diacylglycerol (DAG) formation (13, 14). Inositol trisphosphate then mediates an immediate mobilization of intracellular calcium stores, leading to a acute spike of increase in [Ca2+]i that rapidly (within 1 min) decays to basal (13, 15, 16, 17). This acute increase in [Ca2+]i together with DAG induce an acute burst of PRL release (15, 16, 18). This is followed by a sustained increase in [Ca2+]i that is due in part to calcium influx via L-type calcium channels (19), and correlates with a sustained increase in DAG. Activation of PKC by DAG may mediate the sustained activation of calcium channels. In addition, PKC activates MAPKs, which are both implicated in TRH-induced PRL secretion. Thus, the mechanism of TRH-induced PRL secretion is complex and appears to recruit a variety of pathways and second messengers.

The key trigger for TRH action appears to be the initiation of phosphatidyl inositol turnover by Gq/11-mediated activation of PLC. Thus, the most direct mechanism for inhibition of TRH action by dopamine would be direct inhibition of PLC activation. However, several previous studies have indicated that dopamine-D2 receptors do not inhibit the initial responses to TRH (20, 21, 22, 23). For example, acute TRH-induced calcium mobilization or phosphatidyl inositol turnover was not inhibited in lactotrophs, or GH4ZR7 pituitary tumor cells which express the dopamine D2S receptor (24). Similarly, other Gi-coupled receptors such as somatostatin receptors also fail to inhibit the acute phosphatidyl inositol turnover or calcium mobilization induced by TRH (25). However, PLC activation and calcium influx was reduced after 1–2 min during the plateau phase of TRH action. This led to the suggestion that dopamine acts indirectly via a calcium-mediated inhibition of PLC activation. Because PLC is a calcium-dependent enzyme, dopamine-induced inhibition of [Ca2+]i could reduce PLC activity. However, this explanation was not particularly appealing because the reduction in calcium is small (less that 50%), whereas the block in PRL secretion is complete. These findings led to the suggestion that both direct (via inhibition of PLC) and indirect mechanisms (via inhibition of L-type calcium channels) mediated D2-induced inhibition of TRH-mediated PLC activation.

Direct evidence that D2 receptors directly inhibit PLC was not demonstrated for over 10 yr (26). In their recent publication, Rasolonjanahary et al. (27) (this issue) have investigated TRH- and angiotensin-II-stimulated IP production in membranes from pituitary cells. In this in vitro membrane preparation, TRH induced a small (30%) increase in PLC activity that was highly dependent on [Ca2+] and GTP concentration. This response appeared to be due to receptor coupling to G proteins because it was GTP dependent and blocked by anti-G{alpha}q/11 antibody. Dopamine partially inhibited by 50% TRH- or angiotensin-II-stimulated increase in inositol phosphate formation, and PTX or antibodies to G{alpha}i1/2 reversed this effect. The authors argue that inefficient coupling of TRH and dopamine-D2 receptors in vitro compared with intact cells may result from a loss of cytosolic components (e.g. scaffolding proteins, regulator of G protein signaling proteins, etc.). Nevertheless, these results support the earlier contention of a direct inhibition of PLC activation by dopamine-D2 receptor activation (21), which could account for D2-induced inhibition of TRH-induced sustained phase of PLC activation, observed in intact cells. In addition, D2-induced inhibition of TRH-mediated calcium channel activation indirectly inhibits the sustained phase of PLC activation by decreasing [Ca2+]i. However, inhibition by dopamine of TRH-induced PLC activation in vitro does not explain the insensitivity to dopamine of the initial TRH-induced burst in phosphatidyl inositol turnover and calcium mobilization that is observed in intact cells. Although the D2 receptor inhibited "basal" PLC activity in vitro, this required a minimal level of PLC activation by either calcium or Gq/11. In intact cells, the basal activity of PLC is not sensitive to dopamine and appears to require activation (e.g. by TRH) to become sensitive to dopamine. In intact cells, TRH-induced PLC activation is apparently more rapid than the kinetics of D2-mediated PLC inhibition, permitting maximal stimulation of PLC by TRH acutely (within seconds) followed by inhibition by dopamine.

The implications of D2-induced inhibition of PLC as a mechanism to inhibit TRH-induced PRL secretion remain to be clarified. Because initial TRH-induced calcium mobilization and PI turnover are insensitive to dopamine, yet TRH-induced PRL secretion is completely inhibited, some other D2-induced mechanism must contribute inhibition of PRL secretion. For example, TRH activates MAPK via PKC- dependent and PKC-independent actions (28), and dopamine-D2 receptor activation inhibits this response (29, 30, 31, 32, 33). However, inhibition of TRH-induced MAPK activation with PD098059 did not block PRL secretion but inhibits TRH-induced transcriptional activation of the PRL gene (31, 34). Thus, the mechanism by which dopamine-D2 receptor activation blocks TRH-induced PRL secretion remains incompletely understood.

The finding of direct D2-induced inhibition of PLC (27) leads to several testable hypotheses that could have implications for a wide variety of Gi/Go-coupled receptors. The finding that G{alpha}i1/2 antibody blocked the D2-induced inhibition of PLC activation suggests that these G proteins couple to PLC. It would be interesting to test whether Go or Gi3 proteins are also capable of inhibitory coupling to PLC. The mechanism by which Gi proteins couple to PLC is unclear: no direct interactions of G{alpha}i proteins with PLC-ß subtypes have been identified to date, as is observed for several AC subtypes. Thus, a mechanism involving direct binding of G{alpha}i to PLC seems unlikely, although this interaction may be difficult to demonstrate if PLC must be preactivated (e.g. by G{alpha}q/11 or calcium). Alternately, G{alpha}i may modulate the calcium sensitivity of PLC and this could be tested in vitro. Because many receptors couple to Gi/Go proteins, the mechanism by which dopamine-D2 receptors inhibit PLC activity is likely to be shared among other receptors. It will be interesting to test whether other Gi-coupled receptors expressed in pituitary cells (e.g. somatostatin, muscarinic-M4, adenosine, etc.) mediate direct inhibition of PLC.

Inhibitory regulation of PLC by dopamine-D2 receptors, and Gi/Go-coupled receptors in general, appears to be highly cell type-selective. Several Gi/Go-coupled receptors including the D2 receptor stimulate rather than inhibit PLC activity when activated in a variety of mesenchymal or immune cells (17). This pathway is also blocked by PTX and involves mobilization of Gß{gamma}-subunits rather than direct actions of G{alpha}i or G{alpha}o (35, 36). Gß{gamma}-subunits activate certain subtypes of PLC, PLC-ß2, and PLC-ß3, which are highly enriched in mesenchymal cells but are absent or not coupled in pituitary cells (37). Thus, Gi-mediated inhibition of PLC may only be evident in cells that lack Gß{gamma}-regulated forms of PLC, such as pituitary cells or neurons. The findings by Rasolonjanahary et al. (27) represent a novel pathway for negative regulation of PLC activity in pituitary cells. Further studies will be required to address the exact mechanism by which G{alpha}i1/2 mediated inhibition of PLC in pituitary cells because to date no direct interactions between Gi/Go proteins and PLC subtypes have been described (38).


   Acknowledgments
 
I thank Dr. Behzad Banihashemi for critical reading of the manuscript.


   Footnotes
 
P.R.A. is Canadian Institutes of Health Research (CIHR)/Novartis Michael Smith Chair in Neurosciences and is supported by the CIHR, Parkinson’s Society of Canada, and National Cancer Institute of Canada.

Abbreviations: DAG, Diacylglycerol.

Received December 21, 2001.

Accepted for publication December 21, 2001.


   References
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