Sequential alteration of microglia and astrocytes in the rat thalamus following spinal nerve ligation.

BACKGROUND: Spinal reactive astrocytes and microglia are known to participate to the initiation and maintenance of neuropathic pain. However, whether reactive astrocytes and microglia in thalamic nuclei that process sensory-discriminative aspects of pain play a role in pain behavior remains poorly investigated. Therefore, the present study evaluated whether the presence of reactive glia (hypertrophy, increased number and upregulation of glial markers) in the ventral posterolateral thalamic nucleus (VPL) correlates with pain symptoms, 14 and 28 days after unilateral L5/L6 spinal nerve ligation (SNL) in rats. METHODS: Mechanical allodynia and hyperalgesia (von Frey filament stimulation) as well as ambulatory pain (dynamic weight bearing apparatus) were assessed. Levels of nine glial transcripts were determined by quantitative real-time PCR on laser microdissected thalamic nuclei, and levels of proteins were assessed by Western blot. We also studied by immunohistofluorescence the expression of glial markers that label processes (GFAP for astrocytes and iba-1 for microglia) and cell body (S100ß for astrocytes and iba-1 for microglia) and quantified the immunostained surface and the number of astrocytes and microglia (conventional counts and optical dissector method of stereological counting). RESULTS: Differential, time-dependent responses were observed concerning microglia and astrocytes. Specifically, at day 14, iba-1 immunostained area and number of iba-1 immunopositive cells were decreased in the VPL of SNL as compared to naïve rats. By contrast, at day 28, GFAP-immunostained area was increased in the VPL of SNL as compared to naïve rats while number of GFAP/S100ß immunopositive cells remained unchanged. Using quantitative real-time PCR of laser microdissected VPL, we found a sequential increase in mRNA expression of cathepsin S (day 14), fractalkine (day 28), and fractalkine receptor (day 14), three well-known markers of microglial reactivity. Using Western blot, we confirmed an increase in protein expression of fractalkine receptor at day 14. CONCLUSIONS: Our results demonstrate a sequential alteration of microglia and astrocytes in the thalamus of animals with lesioned peripheral nerves. Furthermore, our data report unprecedented concomitant molecular signs of microglial activation and morphological signs of microglial decline in the thalamus of these animals.

Activating PV-positive neurons in ventral thalamic reticular nucleus reduces pain sensitivity in mice.

Previous studies have demonstrated that thalamic reticular nucleus (TRN) and the sub-nuclei play important roles in pain sensation. Our previous findings showed that activating parvalbumin-positive (PV+) neurons in dorsal sector of TRN (dTRN) could reduce the pain threshold and consequently increase the pain sensitivity of mice. Recent studies have shown that activation of GABAergic projection of TRN to ventrobasal thalamus (VB) alleviated pathological pain. GABAergic neurons in TRN are mainly PV+ neurons. However, the exact roles of ventral TRN (vTRN) PV+ neurons in pain sensation remain unclear. In this study, the designer receptors exclusively activated by designer drugs (DREADD) method was used to activate the PV+ neurons in vTRN of PV-Cre transgenic mice, and the mechanical threshold and thermal latency were measured to investigate the regulatory effects of vTRN on pain sensitivity in mice. Thereafter, PV-Cre transgenic mice, conditional anterograde axonal tract tracing, and immunohistochemistry were used to investigate the distribution of PV+ neurons fibers in vTRN. The results showed that the activation of PV+ neurons in vTRN increased the mechanical threshold and thermal latency, which indicated reduction of pain sensitivity. The fibers of these neurons mainly projected to ventral posterolateral thalamic nucleus (VPL), ventral posteromedial thalamic nucleus (VPM), ventrolateral thalamic nucleus (VL), centrolateral thalamic nucleus (CL) and various other brain regions. These findings indicated that activation of PV+ neurons in the vTRN decreased pain sensitivity in mice, which provided additional evidence on the mechanisms of PV+ neurons of TRN in regulating neuralgia.