Mark A. Esrick

A new chemotaxis assay shows the extreme sensitivity of axons to molecular gradients

Axonal chemotaxis is believed to be important in wiring up the developing and regenerating nervous system, but little is known about how axons actually respond to molecular gradients. We report a new quantitative assay that allows the long-term response of axons to gradients of known and controllable shape to be examined in a three-dimensional gel. Using this assay, we show that axons may be natures most-sensitive gradient detectors, but this sensitivity exists only within a narrow range of ligand concentrations. This assay should also be applicable to other biological processes that are controlled by molecular gradients, such as cell migration and morphogenesis.

Non-invasive, in-vivo electrical impedance of EMT-6 tumours during hyperthermia: Correlation with morphology and tumour-growth-delay

The electrical impedance at frequencies from 100 Hz to 40 MHz of EMT-6 tumours was measured non-invasively, in vivo, during hyperthermia using an apparatus constructed for this purpose. Histology and morphometry were performed on tumours harvested periodically during the heating. A ratio of conductivities at two frequencies (sigma (10MHz)/sigma (10kHz)), which minimizes the tissues temperature-coefficient effects, was used to correlate impedance changes with the histopathological changes. The bulk of the cell population followed a necrotic cell death sequence during heating. Initial increase of the sigma-ratio correlated with cell swelling, and a reversal of the rate of this increase correlated with the appearance of small membrane breaks and evidence of mitochondrial damage. A continued, slowing sigma-ratio increase to a maximum correlated with continued cell swelling accompanied by increasing membrane disruption. The subsequent decrease in sigma-ratio correlated with continued general cell lysing. Between the appearance of the first membrane breaks (sigma-ratio peak) and the evidence of general lysing (sigma-ratio peak), the tumour-growth-delay increased non-linearly. Because the sigma-ratio consistently discerned these events, these measurements were able to predict the fate of this cell population when subjected to hyperthermia. Knowledge of temperature or time of heating was not required.

Monitoring temperature and tissue-dependent changes via electrical impedance trajectories during therapy

The low frequency (<EQ 10 MHz) electrical impedance of a volume of tissue is sensitive to its temperature and its response to heating and other stresses. Major tissue changes, such as those accompanying higher hyperthermic temperatures or prolonged ischemia, and not necessarily reversible unless detected in time to alleviate the stress. Thus, it is imperative to assess the temperature and/or tissue changes in real-time if adequate monitoring of thermal treatments is to be accomplished. To this end, we focus on the use of electrical impedance measurements of a volume of tissue at temperatures in the hyperthermia region (<EQ 47 degree(s)C) where tissue responses occur at a rate which is controllable. First, using well controlled freshly excised tissue data, we examine the prototypical impedance changes associated with the early and later stages of necrosis within a tissue subjected to heating and ischemia. Then, impedance measurements made non-invasively, in vivo, in HT29 tumors are used to demonstrate the differences caused by different thermal treatments and from differences in the ischemic condition of the tissues. The electrical impedance signature of the tumors were indicative of certain cellular-level changes occurring within the tumors. The histological findings corroborate the ability of the electrical impedance to report these cellular changes. The changes are consistent with the cells proceeding along the path of necrosis. Initial cell swelling appears to be largely due to ischemia, and the cell lysing and tumor response to the defined period of heating.