Food intake is stimulated following adminstration of orexin-A into the perifornical region of the lateral hypothalamus (LH/PFA). Orexin neurons originating in the LH/PFA interact with a number of hypothalamic systems known to influence food intake, including glutamatergic neurons. Glutamatergic systems in the LH/PFA were demonstrated to initiate feeding through NMDA receptors. Male Sprague-Dawley rats fitted with brain guide cannulae to the LH/PFA were used in two experiments. In one, a combination microdialysis/microinjection probe was used to deliver aCSF or 500 pmoles orexin-A into the LH/PFA. Orexin-A increased interstitial glutamate to 143 ± 12% of baseline (p<0.05), which remained elevated over the 120 min collection period. In the second, the N-methyl-D-aspartic acid (NMDA) receptor antagonist D-2-amino-5-phosphonopentanoic acid (D-AP5) (10 nmoles) was administered prior to orexin-A. The orexin-induced increase in food intake (from 1.1 ± 0.4 to 3.2 ± 0.5 g, p<0.05) during the first hour was absent in rats receiving D-AP5 + orexin-A (1.2 ± 0.5 g). There was no effect of D-AP5 alone on food intake. These data support glutamatergic systems in the LH/PFA mediating the feeding response to orexin-A through NMDA receptors.