The ocular effects of spitting cobras: I. The ringhals cobra (Hemachatus haemachatus) venom-induced corneal opacification syndrome.

Venom from the ringhals cobra (H. haemachatus) caused extensive chemosis, prolonged corneal edema and marked miosis when applied locally to the eyes of two strains of rabbits: Buskat (albino) and Chinchilla (pigmented). In the pigmented rabbits, the corneal edema progressed to complete corneal opacification with corneal and conjunctival neovascularization that remained unchanged until the end of the experiment (70 d). In the albino rabbits the corneal cloudiness and conjunctivitis cleared within three weeks of venom instillation. Treatment with heparin and tetracycline markedly improved the corneal opacification syndrome, while treatment with specific antivenom resulted only in partial improvement. It is postulated that the venom owes its ocular effects to its cardiotoxin, the only venom fraction capable of inducing the corneal opacification syndrome. The protective effect of heparin is probably through its electrostatic binding to the cardiotoxin in the venom. Tetracycline could form ionic bonding at several sites in the strongly basic cardiotoxin. The corneal opacification syndrome was associated with the ability of the snakes to spit since venoms from the other non-spitting cobras were devoid of this activity.

The ocular effects of spitting cobras: II. Evidence that cardiotoxins are responsible for the corneal opacification syndrome.

Fractionation of H. haemachatus, N. nigricollis, N. nivea and N. melanoleuca venoms using Amberlite CG-50 and (NH4)HCO3 elution gradient chromatography yielded 11-13 fractions for each venom. One fraction, F X, from H. haemachatus, two fractions, F X and F XI, from N. nigricollis and one fraction, F VIII, from N. melanoleuca venoms possessed the whole of ocular activity of the venoms. The fractions were the only venom fractions that caused cardiac depressant activity; their effect was reversed by raising Ca++ concentration in the physiological solution; they did not influence the twitches of the phrenic nerve hemidiaphragm and guinea-pig ileum preparations. Further purification of the fractions on Sephadex G-50 followed by fractionation on Amberlite CG-50 yielded fractions free from phospholipase A2 activity but possessing the same ocular effects. Similarly, the cardiotoxin from commercial N. nigricollis venom caused the same ocular effects as the crude venom and its purified cardiotoxic fractions. All cardiotoxic fractions as well as N. nigricollis cardiotoxin, caused extensive chemosis, blepharitis and corneal opacification with corneal and subconjunctival neovascularization. On a weight basis, the cardiotoxins were weaker in their oculotoxic activity than the corresponding parent crude venoms possibly because of the potentiating effect of phospholipase A2 in the crude venoms. It is postulated that in spitting cobras the cardiotoxins are responsible for the corneal opacification syndrome. In other cobra venoms the stable binding of cardiotoxins with acidic proteins limits their possible ocular effects. Only in the venoms of the spitting species are the cardiotoxins present in an appropriately free form to cause the ocular opacification syndrome.