Organ-cultured chick embryonic hearts of various ages. I. Electrophysiology

Tetrodotoxin (TTX)-insensitive slow Na⁺ channels are converted or replaced by TTX-sensitive fast Na⁺ channels during normal embryonic development of the chick heart, and rapid reversion occurs in monolayer cell culture (denervated). Fast Na⁺ channels first appear at 4 to 5 days, which is about the time of innervation. Studies were done to determine whether changes in cation channels will occur while hearts are in organ culture. To test whether fast Na⁺ channels will develop in the absence of innervation, hearts from chick embryos 2 to 3 days old were placed into culture for 6 to 8 days. Although the resting potentials of the ventricular cells were about the same as those obtained from fresh 8 to 10 day old hearts, the maximum rate of rise of the action potentials ($\text{+}\text{V}\text{?}{}_{\mathrm{<mtext>max</mtext>}}$) did not reach the high value (about 80 V/s) expected from the calendar age. Instead $\text{+}\text{V}\text{?}{}_{\mathrm{<mtext>max</mtext>}}$ remained at about the same value (12 V/s) that the hearts had when placed into culture. The action potentials were completely insensitive to TTX. The slow channels admit primarily Na⁺ and not Ca²⁺ because Mn²⁺ (1 mm) and lowering [Ca²⁺]₀ to nearly zero by EGTA did not diminish $\text{+}\text{V}\text{?}{}_{\mathrm{<mtext>max</mtext>}}$. To test whether the fast Na⁺ channels disappear in organ culture, hearts from embryos 15 to 19 days old were cultured as whole hearts or minced hearts. The whole hearts survived well for 1 to 6 days; the $\text{+}\text{V}\text{?}{}_{\mathrm{<mtext>max</mtext>}}$ values remained high (～ 100 V/s), and TTX completely blocked the action potentials. The minced hearts had variable $\text{+}\text{V}\text{?}{}_{\mathrm{<mtext>max</mtext>}}$ values, depending on the piece. Those pieces which had a low $\text{+}\text{V}\text{?}{}_{\mathrm{<mtext>max</mtext>}}$ were insensitive to TTX, and those which had a high or intermediate $\text{+}\text{V}\text{?}{}_{\mathrm{<mtext>max</mtext>}}$, were reduced to 5 to 20 V/s by TTX; these persisting responses in TTX were not blocked by Mn²⁺ or zero [Ca²⁺]₀. The results suggest that, while in organ culture, young hearts do not gain fast Na⁺ channels or lose the slow Na⁺ channels that would normally occur in situ. Organ-cultured old hearts left intact do not lose their fast Na⁺ channels. Thus, young or old hearts retain the channels that they originally possessed when placed into culture. Mincing initiates a gain of slow Na⁺ channels, and in some pieces, a partial loss of fast Na⁺ channels.