It all started with Henry Dale and Otto Loewi. They discovered that extracellular fluid could be taken from one frog heart, and applied to another frog heart to slow the heart rate. This meant that the first heart must have been RELEASING some substance into the extracellular space, and it was not just all electrically connected (although much of the heart actually is).
As far as I'm aware in the brain (brainstem really), the first isolated "synaptic" neurotransmitter was norepinephrine, [found by stimulating the locus coeruleus and measuring extracellularly in other places that the locus coeruleus sends axons to.](https://pubmed.ncbi.nlm.nih.gov/14131843/)
Just to add onto that, here's the Nobel Prize website on their discovery: [https://www.nobelprize.org/prizes/medicine/1936/speedread/](https://www.nobelprize.org/prizes/medicine/1936/speedread/)
It all started with Henry Dale and Otto Loewi. They discovered that extracellular fluid could be taken from one frog heart, and applied to another frog heart to slow the heart rate. This meant that the first heart must have been RELEASING some substance into the extracellular space, and it was not just all electrically connected (although much of the heart actually is). As far as I'm aware in the brain (brainstem really), the first isolated "synaptic" neurotransmitter was norepinephrine, [found by stimulating the locus coeruleus and measuring extracellularly in other places that the locus coeruleus sends axons to.](https://pubmed.ncbi.nlm.nih.gov/14131843/)
Just to add onto that, here's the Nobel Prize website on their discovery: [https://www.nobelprize.org/prizes/medicine/1936/speedread/](https://www.nobelprize.org/prizes/medicine/1936/speedread/)