Indoleamines and calcium enhance somatic embryogenesis in Coffea canephora P ex Fr

Melatonin (MEL) and serotonin (SER) are important indoleamines that are involved in neural transmission in mammalian cells. They are also known to be present in various genera of plants. The role (s) of these indoleamines in plants are not well known. In this study, the effects of SER, MEL, calcium, and calcium ionophore (A23187), a calcium channel activator, on somatic embryogenesis in Coffea canephora have been investigated. Adding 100 μM of either SER or MEL to ½ strength Murashige and Skoog (MS) medium and 0.93 μM kinetin (KN) has resulted in enhanced induction of somatic embryogenesis, 85 ± 3 and 62 ± 6 embryos/callus, respectively. In the presence of either 5 mM calcium or 100 μM calcium ionophore A23187, number of somatic embryos/callus is also increased, with 56 ± 4 and 118 ± 10 somatic embryos/callus, respectively, compared to 25 ± 3 embryos/callus for control. The presence of 5 mM calcium chloride along with either 100 μM SER or 100 μM MEL, respectively, have also promoted somatic embryogenesis with induction of 105 ± 6 and 78 ± 2 somatic embryos/callus. While, addition of calcium ionophore A23187 along with either 100 μM SER or 100 μM MEL have produced 155 ± 12 or 135 ± 8 embryos/callus, respectively. In contrast, addition of such indoleamine inhibitors as 40 μM p-chlorophenylalanine (p-CPA), 20 μM fluoexitine hydrochloride (prozac), 1 mM verapamil hydrochloride (calcium channel blocker), and 1 mM ethylene glycol-bis (β-amino ethylether)-N, N, N′, N′-tetra acetic acid (EGTA) (a calcium chelator) individually, has inhibited induction of somatic embryos while reducing levels of endogenous pools of SER, MEL and indole-3-acetic acid (IAA) levels. Calcium imaging by laser scanning confocal microscopy (LSCM) has revealed high fluorescence intensity in callus treated with calcium and calcium ionophore A23187. Immunolocalization of SER in different tissues of C. canephora has revealed that it is localized in vascular tissues of stems, roots, and somatic embryos, as well as in endocarps (husks) of immature fruits.