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Thyrotropin-releasing Hormone ( TRH ) Receptor Number Determines the Size of the TRH-responsive Phosphoinositide Pool

We use an adenovirus vector, AdCMVmTRHR, to express thyrotropin-releasing hormone (TRH) receptors (TRH-Rs) to determine whether the size of the hormoneresponsive phosphoinositide pool in mammalian cells is directly related to receptor number. Infection of HeLa cells with increasing numbers of AdCMVmTRHR caused time-dependent graded expression of TRH-Rs. Measurement of cytoplasmic free Ca2+ in individual cells permitted quantitation of the fraction of cells responsive to TRH. Infection with 100 AdCMVmTRHR particledcell or more led to TRH responsiveness in 290% of HeLa cells. Measurement of prelabeled phosphoinositides hydrolyzed during prolonged TRH stimulation assesses the size of the TRH-responsive pool. In cells infected with AdCMVmTRHR for 24 h, the size of the TRH-responsive phosphoinositide pool increased with increasing TRH-R expression. The “RH-responsive pool also increased with time after infection as the number of TRH-Rs increased. Similar observations were made in G W and KB cells. These data confirm our previous suggestion (Cubitt, A. B., Geras-Raaka, E., and Gershengorn, M. C. (1990) Biochern. J. 271,331436) that there are hormoneresponsive and -unresponsive pools of cellular phosphoinositides and that the maximal size of the TRHresponsive pool is directly related to the number of TRH-Rs.