It remains unknown what proportion of delayed arterial narrowing after subarachnoid hemorrhage depends on ongoing metabolic activity within arterial smooth muscle cells versus changes in the passive structural properties of the arterial wall. To determine this, vasospasm was induced by the double subarachnoid hemorrhage model. Anterior spinal artery segments were harvested from control dogs and from dogs with vasospasm. The segments were suspended in a force transducer and stretched to an optimal length for contraction. The difference in tension between 37 and 0 degrees C was defined as the intrinsic tone, and the residual tension at 0 degrees C was defined as the passive tension. The segments taken from dogs with vasospasm had increased intrinsic tone and passive tension (the differences were 3.8 kN/m2 [P < 0.05] and 14.8 kN/m2 [P < 0.025], respectively). Hence, the passive component accounted for 79.6% of the increased tension in vasospastic arterial segments. The intracellular calcium concentration was measured in these segments, using the luminescent calcium indicator, aequorin. The vasospastic segments had increased basal intracellular calcium concentration (398 versus 258 nmol/L, P < 0.025). In parallel experiments, control and vasospastic vessels were immediately excised when the animals were killed, and the vessels were quick-frozen. Subsequently, using two-dimensional gel electrophoresis to measure percent myosin light chain phosphorylation, vasospastic vessels were found to have increased myosin light chain phosphorylation (37 versus 2%, P < 0.05). The increased intracellular calcium concentration and increased percent myosin light chain phosphorylation in vasospastic segments implicate a role for the Ca(2+)-dependent pathway of smooth muscle cell contraction in vasospasm.
Cell sorting is the workhorse of biological research and medicine. Cell sorters are commonly used to sort heterogeneous cell populations based on their intrinsic features. Raman-activated cell sorting (RACS) has recently received considerable interest by virtue of its ability to discriminate cells by their intracellular chemical content, in a label-free manner. However, broad deployment of RACS beyond lab-based demonstrations is hindered by a fundamental trade-off between throughput and measurement bandwidth (i.e., cellular information content). Here we overcome this trade-off and demonstrate broadband RACS in the fingerprint region (300 - 1,600 cm-1) with a record high throughput of ~50 cells per second. This represents a 100x throughput increase compared to previous demonstrations of broadband fingerprint-region RACS. To show the utility of our RACS, we demonstrate real-time label-free sorting of microalgal cells based on their accumulation of carotenoids and polysaccharide granules. These results hold promise for medical, biofuel, and bioplastic applications.
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