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Ca2+ handling properties of microsomal subfractions of rat vas deferens smooth muscle
Abstract:
The rat vas deferens smooth muscle microsomes on isopycnic centrifugation gave two fractions, namely F2 (15-30% sucrose) and F3 (30-40% sucrose), with comparable ATP-dependent azide-insensitive Ca2+-uptake capacities, although these fractions differed from each other in various enzyme marker activities. The fractions F2 and F3 also show similar pH profiles for the ATP-independent and ATP-dependent Ca2+ uptake, and similar ionized Ca2+-concentration dependence for the ATP-dependent Ca2+ uptake. However, the fractions F2 and F3 differ from each other in that: (a) F3 shows higher permeability to Ca2+, and (b) F3 shows higher stimulation of the ATP-dependent Ca2+ uptake by oxalate. The F3 fraction can also be used to obtain membrane vesicles loaded with Ca2+ oxalate in the presence of ATP. However, the yield of the Ca2+ oxalate enriched fraction is too low to permit their further characterization.
Insights
Two rat vas deferens smooth muscle microsome fractions, F2 and F3, exhibit similar calcium (Ca2+) uptake but differ in permeability and oxalate stimulation. Fraction F3 shows enhanced Ca2+ permeability and oxalate responsiveness.
Area of Science:
- Cellular and Molecular Biology
- Muscle Physiology
- Biochemistry
Background:
- Smooth muscle microsomes are crucial for understanding cellular calcium (Ca2+) regulation.
- Isopycnic centrifugation is a key technique for isolating subcellular fractions.
- Investigating Ca2+ uptake mechanisms in smooth muscle is vital for physiological studies.
Purpose of the Study:
- To characterize and compare two distinct fractions (F2 and F3) derived from rat vas deferens smooth muscle microsomes.
- To elucidate differences in Ca2+ uptake, permeability, and response to oxalate between these fractions.
- To explore the potential of fraction F3 for generating Ca2+-oxalate loaded vesicles.
Main Methods:
- Isopycnic centrifugation of rat vas deferens smooth muscle microsomes to obtain sucrose density fractions (F2 and F3).
- Assay of ATP-dependent and ATP-independent Ca2+ uptake capacities.
- Enzyme marker activity analysis, pH profile determination, and ionized Ca2+-concentration dependence studies.
Main Results:
- Fractions F2 and F3 displayed comparable ATP-dependent, azide-insensitive Ca2+ uptake capacities.
- Both fractions showed similar pH profiles and ionized Ca2+-concentration dependence for Ca2+ uptake.
- Fraction F3 exhibited higher Ca2+ permeability and greater stimulation of ATP-dependent Ca2+ uptake by oxalate compared to F2.
Conclusions:
- Rat vas deferens smooth muscle microsomes yield distinct fractions (F2, F3) with differential Ca2+ handling properties.
- Fraction F3 possesses unique characteristics, including enhanced Ca2+ permeability and oxalate responsiveness.
- While F3 can form Ca2+-oxalate loaded vesicles, low yield limits further characterization.