Formation constants have been determined for the substitution of co-ordinated H20 in aquocobalamin in aqueous solution (ionic strength usually 0.5) at room temperature by the following ligands : CH3-CO2-, 4-5 I./mole ; N3-, 7.2 x lo4; NO2-, 2.3 x l o 5 ; CH3.NC, 7 x lo4;2.2 x l o 7 ; thiourea, 13. No complex was detected with the ligands CO, O,, NO, (CH,)&, or Ph3P. Additional formation constants relating to the thermodynamic trans-effect have been determined (i) when the trans-ligand is SO3,-, and (ii) for the substitution of H 2 0 by N3-, CH,*NC, and SOS2-with different trans-ligands. It is shown that SO,2-is comparable with H C Xand CH2=CH-in its cis-and trans-effects. The absence of CO complexes and the close parallel between formation constants involving CH3*NC and N3are evidence for the absence of any significant .rr-bonding between the cobalt(ll1) ion and the ligand.

The urinary bladder responds to parasympathetic nerve stimulation by contraction but it has been shown in several species that this response is not blocked by atropine. Acetylcholine applied to isolated bladder preparations, on the other hand, produces a contraction which is abolished by atropine in the manner characteristic of a muscarinic response.As early as 1911, Langley observed this anomalous behaviour of the bladder in the dog, cat and rabbit but until recently studies of this phenomenon in other species have been few. Burnstock & Campbell (1963) and Burnstock, O'Shea & Wood (1963) have described its occurrence in the ring-tailed possum (Pseudocheirusperegrinus) and the toad (Bufo marinus). Carpenter (1963) also noted the same results for the bladder of the rat.This paper describes studies on the response of the isolated bladder of the guinea-pig to electrical stimulation and to application of drugs. METHODSAdult guinea-pigs of either sex, weighing 250 to 400 g, were killed by a blow on the head. Both ureters were tied and cut and the bladder was flushed out with 1 to 2 ml. of Locke solution through a cut in the urethra. A glass cannula was then inserted into the bladder through the urethra, tied into place and filled with Locke solution to distend the bladder. The cannulated bladder was immersed in the organ-bath (25 ml. capacity) filled with Locke solution and bubbled with 95%. oxygen and 5% carbon dioxide. The cannula was connected to a U-tube containing black ink and the intraluminal pressure under these conditions was 60 to 80 mm of water. One arm of the U-tube passed through a close-fitting sleeve in which length-wise slits had been cut on opposite sides. The sleeve was illuminated by a lamp and the light passing through the slit was allowed to fall on a selenium photoelectric cell. Movement of the bladder displaced the fluid in the cannula, varying the level of the ink of the U-tube and so changing the illumination of the photo-cell. The potential changes so produced were recorded directly on a self-balancing potentiometric recorder. For electrical stimulation one platinum electrode was inserted through the cannula into the lumen of the bladder and another, which was made the anode, was placed directly in the organ-bath. The shocks were rectangular, submaximal (2 to 5 V) or supramaximal (20 V), and of 0.5 to 1 msec duration. The stimulus frequency was varied. The preparation showed considerable spontaneous activity, although this could be reduced by keeping the bath temperature at 300 C.Treatment ofguinea-pigs with reserpine. Reserpine was dissolved in a 20% solution of ascorbic acid and a dose of 10 mg/kg was given intraperitoneally on four successive days, the animal being used on the fifth day. During this treatment the animals were kept in a warm room.

Formation constants have been determined for the substitution of co-ordinated H, O in aquocobalamin (vitamin B, , , ) in aqueous solution of ionic strength 0 . 5 ~ (KNO, or NaCIO,) a t room temperature by the following anions :F-just detectable, CI-1.3, Br-1.9. I-32, NCO-530, NCS-1200, NCSe-8300, S203,-7300. The cobaltic ion in the cobalamins therefore has mild class ( b ) character. The cobalamins containing I-, NCSe-, and S203,show anomalous spectra similar to those of the alkylcobalamins. The pK for the protonation and displacement of the co-ordinated benziminazole by H, O varies with the nature of the trans-ligand in the order CI-< Br-, Ior, in general, H,O, NH,, CI-< CN-, carbanions, sulphur-containing ligands. Br-. I-. It is shown that the main effect of the axial ligand, both on the spectrum of the equatorial ring system (cis-effect) and on the equilibrium between H 2 0 and benziminazole (trans-effect), is related to the position of the ligand in the nephelauxetic series.

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We have studied the effect of varying one axial ligand (X). usually an organo-ligand, on the stretching frequency of cyanide co-ordinated in the trans-position (Y) and on the spectrum of the equatorial corrin ring in these cyanocomplexes, and have measured several additional formation constants for substitution of the axial ligands. The cyanide stretching frequency is very dependent on the nature of X, falling from 21 32 cm.-l when X = benzimidazole to 2082 cm.-l when X = CH3CH2-. All the evidence relating to cis-and trans-effects in corrinoids which contain only the light ligand atoms C, N. and 0 is compared. It is shown that there is a correlation between cis-and trans-effects and between ground-state and thermodynamic effects, and that, in general, as the ligand X becomes more polarisable, so the other metal-ligand bond lengths increase, cyanide co-ordinated in the trans-position becomes more ionic, and the formation constants for the substitution of H, O fall to values more typical of ion-pairs. It is concluded that the main factor is the amount of negative charge donated to the cobalt atom through the o-bond.WE have been studying the interactions of ligands in cobalt (111) corrinoids, which are complexes related to vitamin B,, (for a diagram of the structure see Part VI 2).These cis-and trans-effects have been divided into three categories:(1) ground-state effects, e g . , effects on bond lengths and angles, bending and stretching force constants; (2) thermodynamic effects, i.e., equilibria;(3) kinetic effects, eg., rates of ligand substitution. The ease of interpretation clearly falls in the order 1 > 2 > 3, and we have concentrated on obtaining data on the ground-state and thermodynamic effects.I n previous papers 1*3-5 we have studied the effect of changing one axial ligand X t on the equilibrium constants between ligands Y and 2 in the trans-position (thermodynamic trans-effect) and on the spectra of the equatorial corrin ring (cis-effect). In this paper we t The axial ligands are designated as follows. X is the ligand whose effect we are studying. For the ground-state effect Y is the ' probe,' e.g., CN-when we study the dependence of the CN stretching frequency on the nature of X. For the thermodynamic effects Y and 2 are the ligands involved in the equilibrium; Y is the common denominator, if any, e.g., H,O in the aquocobinamides and aquocobalamin, Bz in the cobalamins.The following abbreviations are used: Bz = the chelating heterocyclic base 5,6-dimethylbenzimidazole, present in the nucleoside side-chain of the cobalamins ; DBC, 5'-deoxyadenosylcobalamin ; DMSO, dimethyl sulphoxide. report experiments on the effect of X on (1) the stretching frequency of cyanide co-ordinated in the trans-position Y (ground-state trans-effect), and (2) the spectrum when Y = CN-, together with (3) a few additional stability constants for the substitution of one axial ligand Y by another 2. Taken together with previous formation constants and spectra when Y = H20 and Bz, this provides a large amount of data on the ground...