Alpha-methyltyrosine reduces the acute cardiovascular and behavioral sequelae in a murine model of traumatic brain injury

Abstract

Background: Increased catecholamines contribute to heightened cardiovascular reactivity and behavioral deficits after traumatic brain injury (TBI); adrenergic receptor blockade has limited success in reducing adverse sequelae of TBI. Injury-induced increases in the synthesis of catecholamines could contribute to adverse outcomes in TBI. Inhibition of catecholamine synthesis with alpha-methyltyrosine (αMT) could offer a benefit after TBI.

Methods: Original research trial in mice randomized to αMT (50 mg·kg -1 ·d -1 ) or vehicle for 1 week after TBI induced by controlled cortical impact. Primary outcomes of cardiovascular reactivity and behavioral deficits were assessed after 1 week. Secondary outcomes included blood brain barrier permeability and quantification of gene transcription whose products determine intraneuronal chloride concentrations, the release of catecholamines, and activation of the sympathetic nervous system. These genes were the alpha-2 adrenergic receptor ("Adra2c"), the sodium-potassium-chloride cotransporter ("Nkcc1"), and the potassium chloride cotransporter ("Kcc2"). We also assessed the effect of TBI and αMT on the neuronal chloride/bicarbonate exchanger ("Ae3").

Results: Traumatic brain injury-induced increases in blood pressure and cardiac reactivity were blocked by αMT. Inhibition of catecholamine synthesis decreased blood brain barrier leakage and improved behavioral outcomes after TBI. Traumatic brain injury diminished the transcription of Adra2c and enhanced expression of Nkcc1 while reducing Kcc2 transcription; αMT prevented the induction of the Nkcc1 by TBI without reversing the effects of TBI on Kcc2 expression; αMT also diminished Ae3 transcription.

Conclusion: Traumatic brain injury acutely increases cardiovascular reactivity and induces behavioral deficits in an αMT-sensitive manner, most likely by inducing Nkcc1 gene transcription. Alpha-methyltyrosine may prove salutary in the treatment of TBI by attenuating the enhanced expression of Nkcc1, minimizing blood brain barrier leakage, and diminishing central catecholamine and sympathetic output. We also found an unreported relationship between Kcc2 and the chloride/bicarbonate exchanger, which should be considered in the design of trials planned to manipulate central intraneuronal chloride concentrations following acute brain injury.

Graphical abstract

Figure 1

αMT normalizes blood pressure after TBI. TBI increased mean arterial blood pressure compared with sham-operated animals at 1 week, and this increase was blocked by administration of αMT.

Figure 2

αMT rescues TBI-induced increases in cardiovascular reactivity. Acute TBI increased LV wall shortening fraction (left) and LV ejection fraction (right). Post-TBI treatment with αMT prevented TBI-induced increases in LV fractional shortening and ejection fraction.

Figure 3

αMT improved cerebral edema after TBI. TBI increased the extravasation of fluorescein dye, and αMT significantly reduced the extravasation of fluorescein from the intravascular compartment.

Figure 4

αMT improves cognitive function after TBI. Acute TBI reduced the exploratory behavior of mice as reflected in the time the rodents spent moving during an object recognition test and post-treatment with αMT increased exploration time. TBI did not affect the amount of time mice spent in the open arm, when compared with a covered arm, of an elevated maze. However, post-TBI treatment with αMT tended to increase the relative amount of time the mice spent in the open arm of the maze when compared with sham-operated or TBI mice.

Figure 5

(Left) αMT normalized cortical gene transcription after TBI. As expected, TBI, as with other stressful injuries, diminished the expression of the alpha-2 adrenergic receptor (Adra2c) which normally serves to diminish catecholamine outflow and turnover from central neurons while Nkcc1 gene transcription was dramatically enhanced (right). An increase in Kcc2 gene transcription, which facilitates the exit of chloride from neurons, is highly correlated with increased expression of the chloride/bicarbonate exchanger Ae3 (Slc4a3), which enhances chloride entry. The chloride/bicarbonate exchanger may serve a compensatory function for increases in Kcc2 expression to help maintain intracellular chloride concentrations at a fixed level.