399
Brief Papers
References [11 W. LORENZ, A. DOENICKE, K. MESSMER, H.-J. REIMANN, M. THERMANN, W. LAHN, J. BERR, A. SCHMAL, P. DORMANN, P. REGENFUSS and H. HAMELMANN,Br. J. Anaesth. 48, 151 (1976). [2] W. LORENZ,H. BARTH,M. THERMANN,A. SCHMAL,P. DORMANN and I. NIEMEYER, Hoppe-Seyler's Z. Physiol. Chem. 355, 1097 (1974). [3] W. LORENZ, H.-J. REIMANN, H. BARTrt, J. KUSCHE, R. MEYER, m. DOENICKE and M. HUTZEL, Hoppe-Seyler's Z. Physiol. Chem. 353, 911 (1972).
[41 W. LORENZ, A. DOENICKE, R. MEYER, H.-J. REIMANN, J. KUSCHE, H. BARTH, H. GEESING, M. HUTZEL and B. WEISSENBACHER,Br. J. Anaesth. 44, 355 (1972). [51 W. LORENZ, A. DOENICKE, I. DITTMANN, P. HUG and B. SCHWAP,Z, Anaesthetist 26, 644 (1977). [6] W. LORENZ, M. THERMANN, K. MESSIER, A. SCHMAL, P. DORMANN, J. KUSCHE, H. BARTH, R. TAUBER, M. HUTZEL, G. MANN and R. UHLIG, Agents and Actions 4, 336 (1974).
Concurrent Gas Chromatographic-Mass Spectrometric Determination of Histidine, Histamine and their 1-Methyl Metabolites by N. MAHY,J. TUSSELLand E. GELPi Instituto de Biofisica y Nearobiologia, Apartado 145, Sardanyola (Barcelona), Spain
Recently a new method was described for the G L C MS determination of TMS derivatives of histamine, 1,4methylhistamine, imidazoleacetic acid and 1,4-methylimidazoleacetic acid [1 I. This work has now been extended to histidine and 1,4-methylhistidine. However, as the difficulties related to the gas chromatography of histidine required a different derivatization approach, an attempt was made to set up a perfluoroacilation procedure in line with that used for profiling other biogenic amines in biological samples [2, 31.
derivatives thus obtained are stable for one week at - 4 ~ The mass spectra of pentafluoropropionated histidine and 1,4-methylhistidine show relatively abundant molecular ions at rn/e 403 and m/e 445 respectively, which confirm the incorporation of two PFP groups in both cases. These values can be accounted for by a structural rearrangement on derivatization which also has its counterpart in the corresponding heptafluorobutyryl derivatives. Histamine and 1,4methylhistamine, as expected, give molecular ions at m/e 257 and m/e 271, corresponding to the monoacylated derivatives.
Methods and results For this purpose, histidine, 1,4-methylhistidine, histamine and 1,4-methylhistamine were reacted directly for 3 hours at 70~ with pentafluoropropionic anhydride (PFPA). No previous esterification of the carboxyl group is needed. The reaction mixture, taken to dryness and redissolved in acetonitrile, can be separated on 3% OV-17 (retention indices: 1736, 1583, 1827 and 1763) and 3% OV225 (retention indices: 2355, 2014, 2443 and 2148). The
References [1] N. MAHY and E. GELEi, J. Chromatogr. 130, 237 (1977). [2] J. SEGURA, F. ARTmAS, E. MARTINEZ and E. GELei, Biomed. Mass Spectrometry 3, 91 (1976). [31 E. PERALTA and E. GELPi, Clin. Chim Acta 73, 13 (1976).
Metabolism of Histamine during Anaphylactic Shock in Guinea-Pigs by J. MIEEVA,M.C. SANTAtS,C. FOUSSARD,F. RUFF and J.-L. PARROT D~partement de Physiologie, Facult~ de Medecine Necker-Enfants-Malades, 156 rue de Vaugirard - 75730 Paris C~dex 15
Histamine metabolism has been studied after intravenous (i.v.) or intraperitoneal (i.p.) injections of antigen in actively sensitized guinea-pigs. Histamine has been measured with an automated fluorometric method [1] in blood (Hb), lung (HI) and liver (Hh). Diamine oxidase (DAO) has been measured [2] in plasma, liver and lung as well as N-methyl transferase [31 in lung (MI) and liver (Mh).
Results and discussion As shown in Table 1, after an i.v. shock, there is a rise of Hb, DAOb and DAOh. On the contrary, after i.p. shock Hb decreases with elevated DAOb and DAOh. If an i.p. shock is performed after DAO and M inhibitors, there is still a decreased Hb although DAO and M are practically inactive. Further experiments are in progress to try to explain
400
Brief Papers
Table 1 Blood and tissue diamine oxidase activity in shock. Blood
Plasma
Lung
Liver
Histamine DAO Histamine DAO (~g/1) (nM/mVmin) ~g/g) (nM/g/h) Control Shocked (ovalbumine
454 _+ 36 0,09 + 0.02
N-MT (nM/g/h)
Histamine DAO (~g/g) (nM/g/h)
N-MT (nM/g/h)
35.7-+_3.2 57-+ 11
219-+23
2.8+__0.2 3 7 9 + 4 2
56+
2015 -+ 27 0.63 -+ 0.19
31.3__+11
56+_20
203-+46
5.0__+1.3 555-+95
42-+ 9
7
228 -+ 32 0.24 + 0.02
50.7 _+ 14
62 _+ 18
242 + 26
3.8 _+0.4
503 _+ 30
67 _+_13
209 -+ 56 0.01 • 0.01
25.3_+2.3
12_+ 6
0
3.5_+0.8
19_+ 14
i.v.) Shocked (ovalbumine
i.p.) Shocked (i.p.) + ~-aminoguanidine + amodiaquine
this Hb fall. Either a very low concentration of DAO and/or M are sufficient to destroy Hb; either there is a possible increase of another catabolism of histamine (acetylation for example); or as in rabbit anaphylactic shock [4], there is a trapping of some histamine-containing cells.
References
2+2
[2] M.C. SANTAIS(personal communication). [3] W. LORENZ,H. BARTHand E. WERLE, Naunyn-Schmeidebergs Arch. Pharmac. 267, 421-432 (1970). [4] F. RUFF, M.C. SANTAIS, C. FOUSSARD, B. GOUGET, R. AMARAL-MARQUESand J.L. PAaI~OT, Sciences (1977) (in press).
[11 F. RUFF, A. SAINDELLE,E. DUTRIPONand J.L. PARROT, Nature 214, 279-281 (1967).
The Role of Calcium and Strontium Ions in the Secretion of Histamine from Mast Cells by J.L. MONGAR,J.C. FOREMANand M. HALLETT University College London
Mast cells taken from the peritoneum of the rat can be induced to secrete their histamine by a variety of stimuli: antigen-antibody reactions, dextran or concanavaiin A. The secretion is accompanied by an uptake of 45calcium [1]. The two processes are closely coupled: (1) They are linearly related; 10% histamine secretion corresponds to a calcium uptake of about 0.08 fmol/cell. (2) They have similar time courses; both are complete within 1 rain. (3) They are both similarly dependent upon pH; with a maximum at 7.5. (4) They are both similarly dependent upon antigen concentration over the range 10-s to 10-3. (5) Phosphatidyl serine increases both processes whereas dibutyryl cAMP and theophylline inhibit them. However, the two processes can be uncoupled by metabolic inhibition; the blocking of both glycolysis and oxidative phosphorylation completely abolishes histamine secretion but leaves the majority of calcium uptake intact. Thus the calcium uptake which proceeds in the absence of the discharge of granules cannot be due to the binding of calcium to released granular material [2, 3]. Strontium ions substituted for calcium, their optimum concentration is 10 mM compared with 1 mM for calcium and the total amount of histamine secreted in response to an external stimulus may be several times greater [4].
Strontium ions also have the property of releasing histamine by a process that occurs in the absence of an external stimulus. The rate of release is slow, reaching a maximum only after more than 240 rain; the accompanying histamine release is also slow. Surprisingly, these slow processes have a similar coupling ratio to that for stimulated cells: in each case a 10% histamine secretion is associated with a strontium uptake of about 0.1 fmol/cell [3]. Unlike the strontium uptake that occurs in stimulated ceils, the spontaneous uptake of strontium is not readily inhibited by dibutyryl cAMP suggesting entry via a route distinct from the physiological calcium channels. Nevertheless, metabolic inhibition is readily achieved indicating that the final stages of the secretory process are the same for the two routes of entry of the divalent ion.
References [11 J.C. FOREMAN, J.L. MONGAR and B.D. GOMPERTS Nature 245, 249-251 (1973). [21 J.C. FOREMAN, M.B. HALLEXX and J.L. MONGAR, J Physiol. (1977) (in press). [31 J.C. FOREMAN,M.B. HALLETT and J.L. MONGAR, Br. J Pharmac. 55,283P-284P (1975). [4] J.C. FOREMANand J.L. MONGA~ J. Physiol. 224, 753769 (1972).
Brief Papers
References [11 W. LORENZ, A. DOENICKE, K. MESSMER, H.-J. REIMANN, M. THERMANN, W. LAHN, J. BERR, A. SCHMAL, P. DORMANN, P. REGENFUSS and H. HAMELMANN,Br. J. Anaesth. 48, 151 (1976). [2] W. LORENZ,H. BARTH,M. THERMANN,A. SCHMAL,P. DORMANN and I. NIEMEYER, Hoppe-Seyler's Z. Physiol. Chem. 355, 1097 (1974). [3] W. LORENZ, H.-J. REIMANN, H. BARTrt, J. KUSCHE, R. MEYER, m. DOENICKE and M. HUTZEL, Hoppe-Seyler's Z. Physiol. Chem. 353, 911 (1972).
[41 W. LORENZ, A. DOENICKE, R. MEYER, H.-J. REIMANN, J. KUSCHE, H. BARTH, H. GEESING, M. HUTZEL and B. WEISSENBACHER,Br. J. Anaesth. 44, 355 (1972). [51 W. LORENZ, A. DOENICKE, I. DITTMANN, P. HUG and B. SCHWAP,Z, Anaesthetist 26, 644 (1977). [6] W. LORENZ, M. THERMANN, K. MESSIER, A. SCHMAL, P. DORMANN, J. KUSCHE, H. BARTH, R. TAUBER, M. HUTZEL, G. MANN and R. UHLIG, Agents and Actions 4, 336 (1974).
Concurrent Gas Chromatographic-Mass Spectrometric Determination of Histidine, Histamine and their 1-Methyl Metabolites by N. MAHY,J. TUSSELLand E. GELPi Instituto de Biofisica y Nearobiologia, Apartado 145, Sardanyola (Barcelona), Spain
Recently a new method was described for the G L C MS determination of TMS derivatives of histamine, 1,4methylhistamine, imidazoleacetic acid and 1,4-methylimidazoleacetic acid [1 I. This work has now been extended to histidine and 1,4-methylhistidine. However, as the difficulties related to the gas chromatography of histidine required a different derivatization approach, an attempt was made to set up a perfluoroacilation procedure in line with that used for profiling other biogenic amines in biological samples [2, 31.
derivatives thus obtained are stable for one week at - 4 ~ The mass spectra of pentafluoropropionated histidine and 1,4-methylhistidine show relatively abundant molecular ions at rn/e 403 and m/e 445 respectively, which confirm the incorporation of two PFP groups in both cases. These values can be accounted for by a structural rearrangement on derivatization which also has its counterpart in the corresponding heptafluorobutyryl derivatives. Histamine and 1,4methylhistamine, as expected, give molecular ions at m/e 257 and m/e 271, corresponding to the monoacylated derivatives.
Methods and results For this purpose, histidine, 1,4-methylhistidine, histamine and 1,4-methylhistamine were reacted directly for 3 hours at 70~ with pentafluoropropionic anhydride (PFPA). No previous esterification of the carboxyl group is needed. The reaction mixture, taken to dryness and redissolved in acetonitrile, can be separated on 3% OV-17 (retention indices: 1736, 1583, 1827 and 1763) and 3% OV225 (retention indices: 2355, 2014, 2443 and 2148). The
References [1] N. MAHY and E. GELEi, J. Chromatogr. 130, 237 (1977). [2] J. SEGURA, F. ARTmAS, E. MARTINEZ and E. GELei, Biomed. Mass Spectrometry 3, 91 (1976). [31 E. PERALTA and E. GELPi, Clin. Chim Acta 73, 13 (1976).
Metabolism of Histamine during Anaphylactic Shock in Guinea-Pigs by J. MIEEVA,M.C. SANTAtS,C. FOUSSARD,F. RUFF and J.-L. PARROT D~partement de Physiologie, Facult~ de Medecine Necker-Enfants-Malades, 156 rue de Vaugirard - 75730 Paris C~dex 15
Histamine metabolism has been studied after intravenous (i.v.) or intraperitoneal (i.p.) injections of antigen in actively sensitized guinea-pigs. Histamine has been measured with an automated fluorometric method [1] in blood (Hb), lung (HI) and liver (Hh). Diamine oxidase (DAO) has been measured [2] in plasma, liver and lung as well as N-methyl transferase [31 in lung (MI) and liver (Mh).
Results and discussion As shown in Table 1, after an i.v. shock, there is a rise of Hb, DAOb and DAOh. On the contrary, after i.p. shock Hb decreases with elevated DAOb and DAOh. If an i.p. shock is performed after DAO and M inhibitors, there is still a decreased Hb although DAO and M are practically inactive. Further experiments are in progress to try to explain
400
Brief Papers
Table 1 Blood and tissue diamine oxidase activity in shock. Blood
Plasma
Lung
Liver
Histamine DAO Histamine DAO (~g/1) (nM/mVmin) ~g/g) (nM/g/h) Control Shocked (ovalbumine
454 _+ 36 0,09 + 0.02
N-MT (nM/g/h)
Histamine DAO (~g/g) (nM/g/h)
N-MT (nM/g/h)
35.7-+_3.2 57-+ 11
219-+23
2.8+__0.2 3 7 9 + 4 2
56+
2015 -+ 27 0.63 -+ 0.19
31.3__+11
56+_20
203-+46
5.0__+1.3 555-+95
42-+ 9
7
228 -+ 32 0.24 + 0.02
50.7 _+ 14
62 _+ 18
242 + 26
3.8 _+0.4
503 _+ 30
67 _+_13
209 -+ 56 0.01 • 0.01
25.3_+2.3
12_+ 6
0
3.5_+0.8
19_+ 14
i.v.) Shocked (ovalbumine
i.p.) Shocked (i.p.) + ~-aminoguanidine + amodiaquine
this Hb fall. Either a very low concentration of DAO and/or M are sufficient to destroy Hb; either there is a possible increase of another catabolism of histamine (acetylation for example); or as in rabbit anaphylactic shock [4], there is a trapping of some histamine-containing cells.
References
2+2
[2] M.C. SANTAIS(personal communication). [3] W. LORENZ,H. BARTHand E. WERLE, Naunyn-Schmeidebergs Arch. Pharmac. 267, 421-432 (1970). [4] F. RUFF, M.C. SANTAIS, C. FOUSSARD, B. GOUGET, R. AMARAL-MARQUESand J.L. PAaI~OT, Sciences (1977) (in press).
[11 F. RUFF, A. SAINDELLE,E. DUTRIPONand J.L. PARROT, Nature 214, 279-281 (1967).
The Role of Calcium and Strontium Ions in the Secretion of Histamine from Mast Cells by J.L. MONGAR,J.C. FOREMANand M. HALLETT University College London
Mast cells taken from the peritoneum of the rat can be induced to secrete their histamine by a variety of stimuli: antigen-antibody reactions, dextran or concanavaiin A. The secretion is accompanied by an uptake of 45calcium [1]. The two processes are closely coupled: (1) They are linearly related; 10% histamine secretion corresponds to a calcium uptake of about 0.08 fmol/cell. (2) They have similar time courses; both are complete within 1 rain. (3) They are both similarly dependent upon pH; with a maximum at 7.5. (4) They are both similarly dependent upon antigen concentration over the range 10-s to 10-3. (5) Phosphatidyl serine increases both processes whereas dibutyryl cAMP and theophylline inhibit them. However, the two processes can be uncoupled by metabolic inhibition; the blocking of both glycolysis and oxidative phosphorylation completely abolishes histamine secretion but leaves the majority of calcium uptake intact. Thus the calcium uptake which proceeds in the absence of the discharge of granules cannot be due to the binding of calcium to released granular material [2, 3]. Strontium ions substituted for calcium, their optimum concentration is 10 mM compared with 1 mM for calcium and the total amount of histamine secreted in response to an external stimulus may be several times greater [4].
Strontium ions also have the property of releasing histamine by a process that occurs in the absence of an external stimulus. The rate of release is slow, reaching a maximum only after more than 240 rain; the accompanying histamine release is also slow. Surprisingly, these slow processes have a similar coupling ratio to that for stimulated cells: in each case a 10% histamine secretion is associated with a strontium uptake of about 0.1 fmol/cell [3]. Unlike the strontium uptake that occurs in stimulated ceils, the spontaneous uptake of strontium is not readily inhibited by dibutyryl cAMP suggesting entry via a route distinct from the physiological calcium channels. Nevertheless, metabolic inhibition is readily achieved indicating that the final stages of the secretory process are the same for the two routes of entry of the divalent ion.
References [11 J.C. FOREMAN, J.L. MONGAR and B.D. GOMPERTS Nature 245, 249-251 (1973). [21 J.C. FOREMAN, M.B. HALLEXX and J.L. MONGAR, J Physiol. (1977) (in press). [31 J.C. FOREMAN,M.B. HALLETT and J.L. MONGAR, Br. J Pharmac. 55,283P-284P (1975). [4] J.C. FOREMANand J.L. MONGA~ J. Physiol. 224, 753769 (1972).
