The pharmaceutical intervention on the interaction between intracellular signaling pathways mediated by Ca²⁺ and cAMP (Ca²⁺/cAMP signaling interaction) could bring important benefits for combating neurodegeneration and diseases related to aging. This discovery emerged from classical neurotransmission studies using rodent vas deferens as a model. From classical reports using this model, the concept of Ca²⁺-dependent processes involved in the neurotransmission (Ca²⁺ influx triggers muscle contraction and neurotransmitter release) is amply accepted. Thus, Ca²⁺ channel blockers (CCB) due to reduction of Ca²⁺ influx through L-type voltage-activated Ca²⁺ channels (VACC) should reduce neurotransmission. Nonetheless, using this model, some studies performed since 1975 reported that reduction of Ca²⁺ influx by low concentrations of CCB (verapamil, diltiazem or nifedipine) produced a paradoxical increase of the contractions mediated by sympathetic nerves, a phenomenon known as “calcium paradox”. Recent studies using adrenal chromaffin cells have also demonstrated that CCB caused a paradoxical increase of the catecholamine release. Because these compounds are blocking the L-type VACC, an augmented nerve-mediated response due to increased neurotransmitter release was an unexpected outcome. In 2013, we revealed that the Ca²⁺/cAMP signaling interaction could properly explain the so-called “calcium paradox”. The original paper published by us in Cell Calcium (2013) has appeared four times in ScienceDirect TOP 25 Hottest Articles lists. In conclusion, these findings may significantly impact on neurodegenerative diseases, thus may stimulate the development of new pharmacological strategies to combat the diseases related to aging.

From basic science, we know that in mammals, increases of the concentration of free Ca²⁺ ions in the cytosol ([Ca²⁺]_c) serve as a messenger signal to couple the stimulus to muscle contraction, or to neurotransmitter release, among other physiological responses 12. A huge number of experiments performed since the discovery of the role of Ca²⁺ in the control of the heart beat 3 have set the dogma that in excitable cells, the increased Ca²⁺ influx by voltage-activated Ca²⁺ channels (VACC) elicited by depolarising stimuli, triggers muscle contraction and the release of neurotransmitters, and hormones. Conversely, the mitigation of Ca²⁺ influx produced by VACC blockers causes a reduction of those responses 45.

The above concepts imply that the enhanced Ca²⁺ entry during cell depolarisation and/or enhanced Ca²⁺ release from the sarco-endoplasmic reticulum (ER) augments the [Ca²⁺]_c and the triggering of the contractile, or secretory responses. However, about four decades ago, a study showed that verapamil at low concentrations produced a paradoxical increase of the contractions mediated by sympathetic nerves from vas deferens 6. On the other hand, nifedipine was recently found to paradoxically augment the exocytosis of catecholamine triggered by double-pulse depolarisations from voltage-clamped bovine adrenal chromaffin cells, another interesting model to study sympathetic neurotransmission 7. How these two L-type VACC blockers can enhance, instead of reducing, the Ca²⁺-dependent contractile and secretory responses? We properly gave a response to this “calcium paradox” in 2013 through the Ca²⁺/cAMP signaling interaction 8.

In the vas deferens, both release and postsynaptic actions of noradrenaline (NA), and other neurotransmitters such as adenosine 5´ triphosphate (ATP), depend on Ca²⁺ influx by VACC, and the ensuing elevations of [Ca²⁺]_c 9. Hence, some authors found that verapamil abolished both noradrenergic and purinergic components of the sympathetic neurotransmission in the vas deferens 1011. In an old report, however, it was showed that verapamil inhibited the sympathetic contractions of the rat smooth muscles (vas deferens), as predictable; nevertheless, this report also described that the low concentrations of verapamil produced a surprising increase of those neurogenic contractions 6. This paradoxical effect was corroborated in 1981 by French and Scott 12, also in these contractions. Furthermore, six years later a third study reported that verapamil and diltiazem increased these sympathetic contractions; this result was attributed to an agonist effect of CCB on L-type VACC, thus increasing Ca²⁺ influx and neurotransmitter release 13. Another published report (two years later) revealed that both, L- type VACC blockers and activator BAY K 8644, elicited similar increases of the neurogenic contractions of the smooth muscles (vas deferens) mediated by sympathetic nerves; the authors did not provide a reasonable explanation for such paradoxical observation 14.

In a study from our laboratory, we could replicate those previous observations in the sympathetically-mediated contractions of the rat vas deferens: low verapamil concentrations produced a small increase in sympathetic contractions, while high CCB concentrations caused full inhibition of these contractions 8. Similar to the effects observed with high concentrations of CCB, various cAMP enhancers (e.g., the phosphodiesterase inhibitor rolipram) and adenylyl cyclase (AC) activators (e.g., forskolin) also reduced sympathetic contractions; however, low verapamil concentrations in the presence of cAMP enhancers caused a significant increase of these contractions. The increased neurogenic contraction can in turn be reduced by adding an AC inhibitor, SQ22536, to lower the cytosolic cAMP level. These findings suggest that interactions in the Ca²⁺/cAMP signaling pathways could play a key role in explaining the "calcium paradox" as observed in the presence of CCB and cAMP enhancers 8. Thus, these findings can dramatically impact on the cardiovascular, neurodegenerative disorders, cancer and other diseases related to aging 151617.

Based on these findings, we have anticipated that the pharmacological modulation of the Ca²⁺/cAMP signaling interaction by combined use of the L-type CCB and cAMP-enhancer compounds could be a novel therapeutic goal for increasing neurotransmission in neurological, and psychiatric disorders, resulted from neurotransmitter release deficit, and neuronal death 1516. This neuroprotector strategy opens a novel pathway for the drug development which is more efficient and safe for the therapy of several neurodegenerative diseases, including Alzheimer´s, Parkinson´s, amyotrophic lateral sclerosis and Huntington´s diseases 1516. Briefly, the reduction of Ca²⁺ influx through L-type VACC produced by CCB enhances the adenylyl cyclase activity (and consequently elevating cAMP levels, please see figure 1). These CCB-effects can be potentiated by cAMP-enhancer compounds (like PDEs inhibitors). In fact, the fundamental mechanisms by which Ca²⁺/cAMP signaling interaction may increase the transmitter release are due: increasing the content of transmitter in the secretory vesicles (please see figure 1) and enhancing the rate of transmitter release. Thus, by elevating cAMP levels, this second messenger may enhance the release of Ca²⁺ from ER. Indeed, Ca²⁺ is crucial for the transmitter release, participating in virtually all the previous mentioned processes: content of transmitter in the secretory vesicles and rate of transmitter release.

In addition, many reports have shown that increase of cytosolic cAMP concentration stimulates neuroprotective effects 1819. Thus, elevating cAMP levels by handling Ca²⁺/cAMP signaling interaction could reduce neuronal death triggered by cytosolic Ca²⁺ overload 1516. Then, the pharmacological handling of the Ca²⁺/cAMP signalling interaction produced by combination of the L-type CCB prescribed in the antihypertensive therapy, and cAMP-enhancer compounds prescribed in the anti-depressive therapy such as rolipram, could be a novel pharmacological strategy for enhancing neurotransmission in neurological, and psychiatric disorders, resulting of neurotransmitter release deficit, and neuronal death 1516. Figure 1 illustrates how the pharmacological modulation of the Ca²⁺/cAMP signaling interaction could be used in human therapy.

In fact, it was demonstrated that the prescription of L-type CCB reduces motor symptoms, and reduces progressive neuronal death in animal model of Parkinson´s disease, indicating that L-type CCB are potentially viable neuroprotective pharmaceuticals 20. Intriguingly, a 1-decade study involving thousands senile hypertensive patients demonstrated that prescription of L-type CCB reduced blood pressure, and risk of dementia, in hypertensive patients, indicating that these pharmaceuticals could be clinically used to treat neurodegenerative diseases 21. These results for the neuroprotective effects of CCB have been reinvestigated in thousands elderly hypertensive patients with memory dysfunction 22. These studies concluded that patients who have taken CCB had their risk of cognitive dysfunction decreased, such as Alzheimer´s disease 22. These findings reinforce the idea that reduction of cytosolic Ca²⁺ overload produced by L-type CCB due to blockade of Ca²⁺ influx could be an alternative pharmacological goal to reduce, or prevent, neuronal death in neurodegenerative diseases.

Due to involvement of the Ca²⁺ and cAMP signaling pathways in the regulation of the cellular differentiation process, it is possible that the pharmacological modulation of the Ca²⁺/cAMP signaling interaction could stimulate the cellular differentiation in stem cells 1516. In this case, this pharmacological strategy could be used in the cell therapy for the treatment of neurodegenerative diseases.

In conclusion, the Ca²⁺/cAMP signaling interaction may dramatically impact on medical research and therapeutics, stimulating the development of new pharmacological strategies for the therapy of human diseases, including: neurodegenerative diseases, and yet cellular therapy using stem cells for combating diseases related to aging.

Caricati-Neto and Bergantin thank the continued financial support from CAPES, CNPq and FAPESP (Bergantin´s Postdoctoral Fellowship FAPESP #2014/10274-3).