The significance of physiological [Ca2+] and [Mg2+] for in vitro experiments on synaptic transmission

Since 1932 $\text{in}$$\text{vitro}$ physiological and pharmacological studies on neuromuscular and other types of synaptic transmission have been carried out usually in Krebs' of similar balanced electrolyte solutions. It has been disregarded, however, that although the total calcium Ca_t] (2.5 mM) and Mg_t] (1.2 mM), are about the same in human plasma and in Krebs' solution, the physiologically important Ca²⁺] and Mg²⁺], primarily because of binding to plasma proteins, are much lower in plasma (1.1 and 0.6 mM) than in Krebs' solution (2.0 and 1.1 mM). We observed that in a modified Krebs' solution in which the Ca_t] and Mg_t] are 1.4 and 0.9 mM respectively and the Ca²⁺] and Mg²⁺] are about the same as in human plasma, the Ca²⁺ dependent volley output of acetylcholine is less and the inhibition of the electrically induced isometric twitch tension of the rat phrenic nerve - hemidiaphragm preparation by nondepolarizing neuromuscular blocking agents and certain antibiotics is greater than in conventional Krebs' solution, in which the Ca²⁺] and Mg²⁺] are higher than $\text{in}$$\text{vivo}$. Similarly, during electrical field stimulation of the guinea-pig myenteric plexus - longitudinal muscle preparation volley output of acetylcholine is lower and the inhibition of the isometric contraction of the muscle by normophine is greater in modified than in conventional Krebs' solution. It is suggested that for greater relevance to $\text{in}$$\text{vivo}$ conditions the Ca²⁺] and Mg²⁺] of balanced electrolyte solutions used in in vitro experiments on synaptic transmission should be the same as in human plasma or in the plasma of the species of the experimental animal.