Growth cones, the terminal structures of elongating neurites are important for development and regeneration. The second messenger cAMP regulates both processes. cAMP is also known to induce growth cone turning. By using time-lapse microscopy and release of cAMP from biological inactive caged compounds the directional response of neuronal growth cones was investigated. Intracellular gradients were induced by asymmetric release of cAMP on one side of the growth cone. Using a new method, which released the same amount cAMP independent of the stimulation pattern it was possible to show that pulsed release in contrast to continuous release was more effective in inducing growth cone turning. From experiments with several pharmacological agents it can be concluded that the protein kinase A plays a major role for the cellular responses and that the gradient steepness causes the different reactions. In cooperation with other scientists it was ruled out that the different responses were caused by non-linearities of the caged compound. Furthermore cAMP gradients were visualized directly in HeLa cells expressing cAMP sensitive ion channels as a reporter system. With this cell line it was also possible to show that the stimulation pattern, meaning pulsed and continuous release respectively, influences the gradient shape. This finding was in accordance with published theories and numerical calculations. Here it was shown for the first time that the spatio-temporal properties of the intracellular gradient are crucial for growth cone turning.
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