Effects of exogenous amines on mammalian cells, with particular reference to membrane flow

Abstract

We have reviewed the evidence that amines accumulate in intracellular vesicles of low pH, such as lysosomes and endosomes. There is consequent elevation of intravesicular pH, and inhibition of receptor-ligand dissociation often results from this pH change. We have argued that the capacity for fusion of such vesicles is also reduced by the high pH. We suggest that the variety of effects of amines on membrane flow and macromolecular transport we describe are at least partly due to such reduced fusion (Figs. 1 and 2). We propose that an internal low pH may facilitate heterologous vesicle-vesicle and vesicle-plasma membrane fusion. There is some evidence that clathrin can accelerate phospholipid vesicle fusion in vitro at low pH (Blumenthal et al., 1983) but no direct evidence on the role of intravesicular pH. This idea is consistent not only with the preceding discussion, but also with the fact that the intracellular membrane-bound compartments least involved in fusion events (e.g. mitochondria) are of neutral or alkaline internal pH. Membrane fusion is certainly required for the formation of vesicles at the periphery of the Golgi apparatus, and possibly earlier in the transport and processing of biosynthetic products in the Golgi (Bergeron et al., 1982). Thus the accumulation of amines in the Golgi may be responsible for several effects on the flow of macromolecules along their translocation pathways. The status of the plasma membrane in this view is complex. It might be argued that the pH dictating the fusion step in endocytosis is that of the extracellular fluid, in which case the inhibitory effects of amines on this process are not explained. However, the rapidity of acidification of the newly formed endocytic vesicles allows the possibility that plasma membrane invaginations might temporarily sequester areas which are of lower pH than that of the bulk extracellular fluid even before fusion, since the proton pumping enzyme(s) are probably present on the plasma membrane. Were this the case, then an acid pH could again be a factor determining membrane fusion at the plasma membrane. The inhibition of endocytosis by weak bases thus may again reflect elevation of pH in a sequestered compartment. From the data on the dependence of response on the concentration of amines, we anticipate that most responses involving membrane flow will be biphasic, with inhibitory effects at low amine concentration, giving way to stimulatory ones at higher concentrations. We suggest that the reported dichotomy between different amines in intracellular membrane fusion systems (D'Arcy Hart, 1982) may result from this concentration dependence.(ABSTRACT TRUNCATED AT 400 WORDS)

Store-activated Ca2+ inflow in Xenopus laevis oocytes: inhibition by primaquine and evaluation of the role of membrane fusion

“…(4) Although the possibility that primaquine inhibits one or more steps in the pathway of activation of SACCs as well as inhibiting Ca# + flow through the SACC pore cannot be completely eliminated, it is considered unlikely. Moreover the conclusion that primaquine interacts with the SACC protein is consistent with reports that primaquine has a wide range of effects on intracellular processes [16][17][18]20,42,43].…”

“…Those authors concluded that the most likely explanation for the effect of primaquine was inhibition of trafficking and recycling of SACCs, although they could not eliminate a direct inhibition by primaquine of the SACCs. The differences in interpretation of the results obtained for megakaryocytes and oocytes might reflect differences in the mechanisms of activation of the SACCs in the two cell types and the fact that primaquine affects several processes in intact cells [16][17][18]20,42,43].…”

Identification and Characterization of Cathepsin B as the Cellular MARCKS Cleaving Enzyme

“…Proteolysis within lysosomes is dependent on an acidic environment (17,22). Treatment of cells with ammonium chloride has been shown to increase lysosomal pH, resulting in the inhibition of normal lysosomal proteolytic events (22,23). To determine if the generation and/or stability of p40 was pH-dependent, radiolabeled cells were treated with 20 mM NH 4 Cl for different times; detergent extracts of these cells were then prepared and analyzed for the presence of immunoprecipitable p40 (Fig.…”

γ-Aminobutyric Acid Type B Receptors Are Constitutively Internalized via the Clathrin-dependent Pathway and Targeted to Lysosomes for Degradation

“…5B, upper panel). Pretreatment of cells with the lysosomotropic drug chloroquine, which increases endosomal and lysosomal pH and impairs lysosomal function (31,32), completely prevented the loss of staining and led to the co-localization of signals in large intracellular clusters (Fig. 5B, bottom).…”