Abstract
Publisher Summary The secretion of hormones from endocrine cells in the anterior pituitary regulates a variety of physiological functions including reproduction, lactation, growth, and metabolism. However, until recently, cellular mechanisms underlying the stimulus-secretion coupling in these cells have been hard to study for two reasons: (1) it was difficult to isolate specific cell types from the heterogenous population of anterior pituitary cells and (2) the intracellular environment could not be manipulated or monitored accurately. The intracellular environment can be altered by diffusing various activators or inhibitors of second messenger pathways, caged molecules, and ion-selective fluorescent indicator dyes into the cell. A thin glass-bottomed Cunningham chamber can be constructed as follows. A circular hole of approximately 20 mm diameter is drilled in the bottom of a 35-mm plastic culture dish (Falcon). A 25-mm circular glass coverslip is cemented onto the bottom of the culture dish with Sylgard 184 elastomer (Dow Corning). The chamber is rinsed with 100% methanol and then several times with distilled water to remove any grease. To facilitate cell attachment, the bottom of the chamber is coated by adding 1 ml of 1% gelatin solution (Sigma). After 5 min, the solution is removed and the chamber is allowed to dry under UV light.
Published Version
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