GABA Activity Mediating Cytosolic Ca²⁺Rises in Developing Neurons Is Modulated by cAMP-Dependent Signal Transduction

Abstract

In the majority of developing neurons, GABA can exert depolarizing actions, thereby raising neuronal Ca 2ϩ . Ca 2ϩ elevations can have broad consequences during development, inducing gene expression, altering neurite outgrowth and growth cone turning, activating enzyme pathways, and influencing neuronal survival. We used fura-2 and fluo-3 Ca 2ϩ digital imaging to assess the effects of inhibiting or activating the cAMP signal transduction pathway on GABA activity mediating Ca 2ϩ rises during the early stages of in vitro hypothalamic neural development. Our experiments stemmed from the finding that stimulation of transmitter receptors shown to either activate or inhibit adenylyl cyclase activity caused a rapid decrease in Ca 2ϩ rises mediated by synaptically released GABA. Both the adenylyl cyclase activator forskolin and the inhibitor SQ-22,536 reduced the Ca 2ϩ rise elicited by the synaptic release of GABA. Bath application of the membrane-permeable cAMP analogs 8-bromo-cAMP (8-Br-cAMP) or 8-(4-chlorophenylthio)-cAMP (0.2-5 mM) produced a rapid, reversible, dose-dependent inhibition of Ca 2ϩ rises triggered by synaptic GABA release. Potentiation of GABAergic activity mediating Ca 2ϩ rises was observed in some neurons at relatively low concentrations of the membrane-permeable cAMP analogs (20 -50 M). In the presence of tetrodotoxin (TTX), postsynaptic Ca 2ϩ rises triggered by the bath application of GABA were only moderately depressed (13%) by 8-Br-cAMP (1 mM), suggesting that the inhibitory effects of 8-Br-cAMP were largely the result of a presynaptic mechanism.The protein kinase A (PKA) inhibitors H89 and Rp-3Ј,5Ј-cyclic monophosphothioate triethylamine also caused a large reduction (Ͼ70%) in Ca 2ϩ rises triggered by synaptic GABA release. Unlike the short-term depression elicited by activation of the cAMP signal transduction pathway, Ca 2ϩ depression elicited by PKA inhibition persisted for an extended period (Ͼ30 min) after PKA inhibitor washout. Postsynaptic depression of GABA-evoked Ca 2ϩ rises triggered by H89 (in the presence of TTX) recovered rapidly, suggesting that the extended depression observed during synaptic GABA release was largely through a presynaptic mechanism. Long-term Ca 2ϩ modulation by cAMP-regulating hypothalamic peptides may be mediated through a parallel mechanism.Together, these results suggest that GABAergic activity mediating Ca 2ϩ rises is dependent on ongoing PKA activity that is maintained within a narrow zone for GABA to elicit a maximal Ca 2ϩ elevation. Thus, neuromodulator-mediated changes in the cAMP-dependent signal transduction pathway (activation or inhibition) could lead to a substantial decrease in GABAmediated Ca 2ϩ rises during early development.

After Damage of Large Bile Ducts by Gamma-Aminobutyric Acid, Small Ducts Replenish the Biliary Tree by Amplification of Calcium-Dependent Signaling and de Novo Acquisition of Large Cholangiocyte Phenotypes

“…In support of our hypothesis, studies have shown that GABA activity mediating cytosolic Ca 2ϩ increases in developing neurons is triggered by changes (activation or inhibition) in cAMP-dependent signal transduction. 49 The de novo acquisition of proliferative and secretory phenotypes of large cholangiocytes by small cholangiocytes is likely attributable to the IP 3 /Ca 2ϩ -dependent activation of AC8, a Ca 2ϩ -dependent AC isoform that plays a key role in the proliferative and secretory functions of large bile ducts. 17 Nine AC isoforms exist in mammalian cells.…”

At Immature Mossy-Fiber–CA3 Synapses, Correlated Presynaptic and Postsynaptic Activity Persistently Enhances GABA Release and Network Excitability via BDNF and cAMP-Dependent PKA

“…In the present experiments, however, forskolin alone was able to mimic the effects of BDNF, and STD-LTP was prevented when the postsynaptic cell was loaded with the membrane impermeable PKA inhibitor PKI 6 -22. This suggests that cAMPdependent PKA may regulate intracellular calcium rise (Obrietan and van den Pol, 1997;Dunn et al, 2006), which in turn would affect BDNF release from the postsynaptic cell. The precise molecular mechanisms regulating this cascade remain to be elucidated.…”

Alcohol inhibits luteinizing hormone-releasing hormone release by activating the endocannabinoid system

“…Furthermore we found that AEA stimulated GABA release in our experimental conditions. Obrietan and van den Pol (29) suggested that cAMP, the second messenger linked to cannabinoid receptors, could inhibit GABA release. The inhibition of LHRH release by AEA and EtOH from the hypothalamus is mediated in part by CB1-r inhibiting AC.…”