Who invented MNI-glutamate?

MNI-glutamate has been used by several groups for 2-photon uncaging in the past 5 years, here I outline the history of its development, along with background on the origin of caged compounds.

A group of chemists at Oxford University lead by John Baltrop found in 1966 that ortho-nitrobenzyl redox chemistry liberated carboxylates from their esters. This was the first report of a photochemical protecting group; one that has turned out to be so useful to biologists over the the past 28 years. This was started by two groups who independently used this photochemistry to liberate biologically significant molecules inside cells: 1. Engels and Schlegger (J. Med. Chem. 20 (1977) 907-911) photoreleased cAMP in rat glioma cells to induce morphological changes (the photosensitive cAMP was only one of six derivatives of cAMP made for this study, the other phosphate esters were hydrolyzed by intracellular esterases-a technique cleverly exploited later by Roger Tsien with the AM esters of BAPTA Nature (1980) 290: 527-8); and 2. Kaplan and co-workers photoreleased ATP inside red cells and studied potassium efflux generated by the sodium pump (Biochemistry (1978) 17: 1929-1935). The latter study is rightly regarded as the more significant one, as Kaplan's experiment was on a rapid time scale. By the way, it was Kaplan's boss, Joe Hoffman, who dubbed ortho-nitrobenzyl-ATP "caged ATP". The name has stuck ever since, despite the (recent) efforts of organic chemists to give the idea other names, in vain attempts to attract more attention and significance to their own work.

From 1970 a group of chemists lead by Armit Patchornik published a series of pioneering papers on photochemical protecting groups. They introduced the ortho-nitroveratryl or 4,5-dimethoxy-2-nitrobenzyl cage ( J. Am. Chem. Soc. (170) 92: 6333-6335) as an extension of the generic photochemistry of Baltrop's 1966 paper. They also were the first to use nitroaromatic redox chemistry to liberate carboxylates from acylindolines ( J. Am. Chem. Soc. (1976) 98: 843-4; J. Am. Chem. Soc. (1981) 103: 7674-5). These studies are the real inspiration for MNI-glu.

Roger Tsien's group made a nitroindolinyl-BAPTA ( J. Am. Chem. Soc. (1989) 111: 7957-68), hoping to cage the chelator. However, the derivative had poor quantum yield, so nitroindolines were forgotten again. Ten years later the Mill Hill group resurrected the protecting group, making 5-methoxycarbonylmethyl-7-nitroindolinyl-glutamate (which they strangely call "NI-glu"-I guess MCMNI-glu is too long). This very important paper was published online on 24 June 1999 (J. Am. Chem. Soc. (1999) 121: 6508-4). They caged glutamate via the hydrolytically stable indolinyl bond, and so solved one of the central practical problems in this field (namely, spontaneous hydrolysis of caged glutamate). However, the quantum yield and absorption were less than optimal, but as soon as this work appeared I realised that MNI-glutamate would work well for 2-photon uncaging.

The reason for this was that before this paper appeared, I had made an analogous caged glutamate (DMCNB-glu). Using this probe, in collaboration with Kasai's lab, we managed to get the first really decent 2-photon uncaging of glutamate (see picture above). I presented this work in a lecture at the Society for General Physiologists annual conference during the summer of 1999 (J. Gen. Physiol. (1999) 114: 1a). I made MNI-glutamate for the first time on 29th March 2000, about 6 months before the Mill Hill group actually published their synthesis of MNI-glu (Tetrahedron (2000) 56: 8197-8205), and over 18 months before their biological evaluation appeared (J. Neurosci. Meth. (2001) 112: 29-42; online 12 October). However, we wanted to do something really interesting with this caged compound, as we realised that MNI-glu had the potential to revolutionise the field, by making extracellular 2-photon uncaging work well for the first time. Thus, I delayed publishing my synthesis of MNI-glutamate until 22 October 2001 (Nature Neurosci. (2001) 4: 1086-1092-this paper was submitted in June 2001).

So, who invented MNI-glutamate?

The Mill Hill group deserve the credit for resurrecting the nitroindoline protecting group (JACS 1999) and doing nice pharmacology of MNI-glu (JNM 2001). However, the lion's share of the credit must go to the Haruo Kasai and his colleague Masanori Matsuzaki, who got DMCNB-glu, then MNI-glu to work fantastically well for 2-photon uncaging, thus fulfilling Winfried Denk's original idea of 2-photon uncaging microscopy (PNAS (1994) 91: 6629-6633). Since then many groups have published cool papers using 2-photon uncaging of MNI-glu, but would have never attempted it if Masanori hadn't first proved its feasibility.