Ultracytochemical detection of guanylate cyclase C activity in alimentary tract and associated glands of the rat. Influence of pH, ATP and the ions Mg2+ and Mn2+.
Intestinal guanylate cyclase C is activated by guanylin, an endogenous peptide. This activity seems to be modulated by adenine nucleotides, the ions Mg2+ and Mn2+, and pH. In this study, we report an ultracytochemical method for the localization of guanylate cyclase C activity at the electron microscope level. We studied the enzymatic activity in the presence or absence of guanylin and/or ATP, in the presence of the ions Mg2+ or Mn2+, and at different pH levels. The greatest distribution of enzymatic activity was detected in samples incubated at pH 8 and 7.4 in the presence of guanylin, Mg2+ and ATP. Guanylate cyclase C activity was detected at the surface epithelium of stomach and intestine, and in liver, exocrine pancreas and parotid gland. In the intestine, enzymatic activity was more widely distributed in the duodenum than in the jejunum-ileum and colon. In the small intestine, activity was more evident in the upper portion than in the basal portion of the villus. In samples incubated at pH 8 and 7.4 in the absence of ATP, enzymatic activity was detected only in small intestine, liver and exocrine pancreas. Enzymatic activity was present in duodenum incubated at pH 8 and 7.4 in the presence of Mn2+ and in the presence or absence of ATP. No samples incubated in all these experimental conditions but at pH 5 or samples incubated in the presence of guanylin only or in the absence of guanylin, displayed guanylate cyclase C activity. Our results suggest that a complete ultracytochemical detection of guanylate cyclase C activity requires guanylin as stimulator, and incubation in the presence of Mg2+ and ATP at pH 8 and 7.4.
Rambotti
M G
MG
Department of Experimental Medicine and Biochemical Sciences, University of Perugia, Italy.
Giambanco
I
I
Spreca
A
A
D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D046508:Culture Techniques; D004064:Digestive System / Q000201:enzymology* / Q000473:pathology; D006162:Guanylate Cyclase / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007422:Intestines / Q000201:enzymology / Q000473:pathology; D008099:Liver / Q000201:enzymology / Q000473:pathology; D008274:Magnesium / Q000378:metabolism; D008345:Manganese / Q000378:metabolism; D010179:Pancreas / Q000201:enzymology / Q000473:pathology; D010306:Parotid Gland / Q000201:enzymology / Q000473:pathology; D051381:Rats; D000074261:Receptors, Enterotoxin; D050705:Receptors, Guanylate Cyclase-Coupled; D018000:Receptors, Peptide / Q000378:metabolism*; D013270:Stomach / Q000201:enzymology / Q000473:pathology; D013363:Submandibular Gland / Q000201:enzymology / Q000473:pathology
D018000:Receptors, Peptide; D008345:Manganese; D000255:Adenosine Triphosphate; D006162:Guanylate Cyclase; D000074261:Receptors, Enterotoxin; D050705:Receptors, Guanylate Cyclase-Coupled; D008274:Magnesium