Lacrimal secretion stimulants: sigma receptors and drug implications

3H-DTG (1.3-di(2-[5-3H]tolyl)guanidine) or 3H-haloperidol was added to sigma-receptors (25 nM) in the presence of 25 nM spiperone and incubated with increasing concentrations of bromhexine derivatives (phenylalkylamines; 10(-9) to 10(-2)M) in membrane homogenate suspensions. IC50 values for two derivatives ranged from 3.2 to 8.8 nM for both radioligands. A preferred derivative, 7A (N,N'-dimethyl-2-phenyl-ethylamine), yielded an IC50 of 7.8 nM for 3H-haloperidol but showed much less affinity in displacing 3H-DTG (IC50 = 900 nM). Applying the technic of Bromberg [Exp. Eye Res., 40:313-320, 1985], in vitro protein secretion rates were measured following stimulation of either lacrimal gland slices or isolated, intact lacrimocytes with the compounds. In vitro protein secretion rates exhibit a dose-response relationship with increases in protein release up to a concentration of 10(-8) to 10(-4) M for various derivatives of bromhexine and 10(-4) M for carbachol. By means of Schirmer strips, tear fluid was collected over a five minute period at 10 and 60 minutes post-dosing following the topical application (50 microliters) to the right eye of New Zealand white rabbits (n = 20-24) of 7A at various concentrations. Incubation of lacrimocytes with 7A alone (10(-4) M), with haloperidol (10(-4) M) alone or in combination show that 7A is acting as an agonist to stimulate protein release, whereas haloperidol is acting as an antagonist to inhibit release. In vivo protein secretion rates also show a dose-response curve (at both 10 and 60 minutes post-dosing) for 7A that reach a statistically significant maximum in the dosed eye at a concentration of 0.15% w/v. Analysis of protein extracts using size exclusion HPLC shows an increase in secretory proteins, particularly tear-specific prealbumin.