Anterograde transneuronal viral tract tracing reveals central sensory circuits from white adipose tissue

Abstract

The origins of the sympathetic nervous system (SNS) innervation of white adipose tissue (WAT) have been defined using the transneuronal viral retrograde tract tracer, pseudorabies virus. Activation of this SNS innervation is acknowledged as the principal initiator of WAT lipolysis. The central control of WAT lipolysis may require neural feedback to a brain-SNS-WAT circuit via WAT afferents. Indeed, conventional tract tracing studies have demonstrated that peripheral pseudounipolar dorsal root ganglion (DRG) sensory cells innervate WAT. The central nervous system projections of WAT afferents remain uncharted, however, and form the focus of the present study. We used the H129 strain of the herpes simplex virus-1 (HSV-1), an anterograde transneuronal viral tract tracer, to define the afferent circuits projecting from WAT to the central nervous system. Siberian hamster inguinal (IWAT) or epididymal WAT was injected with H129 and the neuraxis processed for HSV-1 immunoreactivity. We found substantial overlap in the pattern of WAT sensory afferent projections with multiple SNS outflow sites along the neuraxis, suggesting the possibility of WAT sensory-SNS circuits that could regulate WAT SNS drive and thereby lipolysis. Previously, we demonstrated that systemic 2-deoxy-d-glucose (2DG) elicited increases in the SNS drive to IWAT. Here, we show that systemic 2DG administration also significantly increases multiunit spike activity arising from decentralized IWAT afferents. Collectively, these data provide structural and functional support for the existence of a sensory WAT pathway to the brain, important in the negative feedback control of lipid mobilization.

Fig. 1.

H129 strain of the herpes simplex virus-1 (HSV-1) injected into inguinal white adipose tissue (IWAT) caused HSV-1 immunoreactivity (-ir) in the ipsilateral dorsal root ganglion (and dorsal horn in the lower thoracic and lumbar spinal cord segments) at 48 h. H129 infection found in the ipsilateral (A) and contralateral (B) dorsal root ganglion (DRGs) at 72 h postinjection was detected using standard immunohistochemistry against HSV-1 with diaminobenzidine peroxidase-generated (brown) chromogen. Sections were then counterstained using cresyl violet (blue).

Fig. 2.

HSV-ir in the thoracolumbar spinal cord dorsal horn 72 h after H129 injections into the ipsilateral inguinal white adipose tissue (A). Ninety-six hours postinoculation, spinal cord H129 infection progressed into the ipsilateral intermediolateral horn (IML) and ventral horn (B). H129 infection was detected using standard immunohistochemistry against HSV-1 (DAB peroxidase-generated chromogen; brown). Sections were counterstained (cresyl violet; blue). Arrows indicate HSV-1-ir fiber from the dorsal horn to the intermedial lateral horn. CC: central canal.

Fig. 3.

H129 injected into inguinal white adipose tissue caused infection in the nodose ganglion, predominantly ipsilateral to the site of injection (A). B: scant H129 infection in the contralateral nodose ganglion is shown for comparison. HSV-ir cells:brown.

Fig. 4.

Low-magnification photomicrograph illustrating HSV-1-ir cells in the gracile nucleus (Gr; A) 114 h after injection into IWAT. B: Gr outlined in A is magnified for better visibility. HSV-ir cells:brown; cresyl violet:blue.

Fig. 5.

H129 injected unilaterally into hamster inguinal white adipose tissue resulted in HSV-ir cells in the brain at 114 h postinjection. A: low-magnification photomicrograph illustrating HSV infection in the brainstem. B: locus coeruleus (LC) and Barrington's nuclei (Bar) outlined in Fig. 5A is magnified for better visibility. C: A5 area outlined in A is magnified. D: raphe (magnus: RMg, pallidus: RPa) regions outlined in A under higher magnification. HSV-ir cells:brown; cresyl violet:blue. 7n, facial nerve, or root; A5, noradrenaline cells; IRt, intermediate reticular nucleus.

Fig. 6.

A: photomicrograph illustrating H129 infection in the caudal midbrain ∼ rostral brainstem 114 h after injection into IWAT. B: Raphe cap (RC)/lateral parabrachial (LPB) region outlined in A is magnified for improved visibility of the H129 infection. C: ventral subcoeruleus nucleus (SubCV) region outlined in A is magnified. HSV-ir cells:brown; cresyl violet:blue. 4n, trochlear nerve, or root; Aq, aqueduct; ml, medial lemniscus.

Fig. 7.

A: photomicrograph illustrating H129 infection in the forebrain at 114 h after inoculation unilaterally into the hamster IWAT. B: portion of the paraventricular hypothalamus (PVH) outlined in A is magnified. HSV-ir cells:brown; cresyl violet:blue. OC, optic chiasm; Re, reuniens thalamus; SCH, suprachiasmatic hypothalamus; ZI, zona incerta.

Fig. 8.

Effects of intraperitoneal administration of 500 mg/kg of 2-deoxy-d-glucose. Intraperitoneal saline is without effect; n = 4. Ketamine/xylazine anesthetized nonfasted male Siberian hamster with right inguinal fat nerve exposed, and isolated in situ inguinal fat pad. Inguinal fat sensory fiber bundles were isolated and transected, and the distal endings, still attached to adipose tissue, were placed on paired Teflon-insulated tungsten electrodes for bipolar multiunit neurophysiological action potential recordings from sensory afferents arising from inguinal fat depot. Representative tracings from one animal is shown.