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. 2024 Dec 24;10(1):e1230.
doi: 10.1097/PR9.0000000000001230. eCollection 2025 Feb.

What do ultrasound vocalizations really mean in rats with different origins of pain?

Yang Yu  1 , Chun-Li Li  1 , Rui Du  1 , Xiao-Liang Wang  1 , Jun Chen  1   2
Affiliations

What do ultrasound vocalizations really mean in rats with different origins of pain?

Yang Yu et al. Pain Rep. .

Abstract

Objectives: This study is to assess how 22 kHz and 50 kHz spontaneous ultrasound vocalization (USV) calls would be affected by different origins of pain so as to validate the use of USV in pain studies.

Methods: Five well-established rat models of pain were used to evaluate various parameters of spontaneous 22 kHz and 50 kHz calls in adult male rats in terms of both acute and chronic or inflammatory and neuropathic or somatic and visceral origins. The effects of local lidocaine blockade of the injection site and intraperitoneal administration of antidepressant (amitriptyline) and anticonvulsant (gabapentin) were examined as well in typical inflammatory and neuropathic pain models, respectively.

Results: The major new gains were as follows: (1) naive rats staying alone and engaging dyadic social interaction with a naive or a conspecific in pain emitted similar power and amounts of both 22 kHz and 50 kHz spontaneous USV calls; however, rats suffering from various origins of pain emitted significantly less USV calls of both 22 kHz and 50 kHz in terms of both number and time. (2) Local blockade of the injury sites of inflammatory pain could reverse the impaired emission of both 22 kHz and 50 kHz spontaneous calls, so did the treatment of the rats with neuropathic pain by amitriptyline and gabapentin.

Conclusions: Emissions of both 22 kHz and 50 kHz spontaneous calls were impaired by acute and chronic pain conditions regardless of inflammatory and neuropathic or somatic and visceral origins.

Keywords: Amitriptyline; Gabapentin; Lidocaine; Pain models; Rat; Ultrasound vocalization.

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Conflict of interest statement

The authors have no conflict of interest to declare.Sponsorships or competing interests that may be relevant to content are disclosed at the end of this article.

Figures

Figure 1.
Figure 1.
Ultrasonographic signal examples of and the effects of social stimulation and pain on powers of spontaneous USVs calls (dB) in adult male rats. (A) Examples of ultrasonographic signals of 22 kHz and 50 kHz calls. (B-D) changes in powers of 22 kHz and 50 kHz calls during 30-minute recordings from rats staying alone (SingleNaive) and those engaging dyadic social interaction (DSI) with a naive (DSINaive-Naive) or a conspecific in pain (DSINaive-Pain). (E-K) Changes in powers of 22 kHz and 50 kHz calls during 30-minute recordings from rats after receiving subcutaneous injection of bee venom (BV), formalin (F), acetic acid (AA), and complete Freund adjuvant (CFA) or spared nerve injury (SNI) preparation. One dot represents 1 ultrasonographic signal seen in (A); n = 8 animals for each single group and n = 6 pairs of rats for DSI. CFA-0.5h and CFA-24h, 0.5 hours and 24 hours after CFA injection; SNI-14d and SNI-30d, 14 and 30 days after SNI preparation; USV, ultrasound vocalization.
Figure 2.
Figure 2.
Averaged powers of spontaneous USV calls in single naive, DSI, and pain-suffering rats. (A) Comparisons of USV powers between SingleNaive and DSINaive-Naive and DSINaive-Pain groups. (B, C) Comparisons of USV powers between single naive rats and rats after receiving subcutaneous injection of bee venom (BV), formalin (F), acetic acid (AA), and complete Freund adjuvant (CFA) or spared nerve injury (SNI) preparation. n = 8 animals for each single group and n = 6 pairs of rats for DSI. *P < 0.05, **P < 0.01 vs single naive group. CFA-0.5h and CFA-24h, 0.5 hours and 24 hours after CFA injection; DSI, dyadic social interaction; SNI-14d and SNI-30d, 14 and 30 days after SNI preparation; USV, ultrasound vocalization.
Figure 3.
Figure 3.
Comparative analysis of USV calls between single naive and DSI groups and the time course effects of pain on emission of USV calls. (A-B) Comparisons of averaged total number and time of 22 kHz and 50 kHz spontaneous calls between SingleNaive, DSINaive-Naive, and DSINaive-Pain groups. (C-D) Time courses of 22 kHz and 50 kHz spontaneous call rate emitted from single naive rats and those received different pain treatments. Data expressed as mean ± SEM; n = 8 animals in each single group and n = 6 pairs of rats for DSI. CFA-0.5h and CFA-24h, 0.5 hours and 24 hours after CFA injection; DSI, dyadic social interaction; SNI-14d and SNI-30d, 14 and 30 days after SNI preparation; USV, ultrasound vocalization.
Figure 4.
Figure 4.
Comparative analysis of USV calls between single naive rats and those received different pain treatments. (A-D) Comparisons of the total number and time of 22 kHz and 50 kHz calls between single naive rats and those after receiving injection of BV, F, AA, and CFA or SNI preparation. Data expressed as mean ± SEM; n = 8 animals in each group; *P < 0.05, **P < 0.01, ***P < 0.001 vs Naive rats. AA, acetic acid; BV, bee venom; CFA, complete Freund's adjuvant; CFA-0.5h and CFA-24h, 0.5 hours and 24 hours after CFA injection; SNI-14d and SNI-30d, 14 and 30 days after SNI preparation; USV, ultrasound vocalization.
Figure 5.
Figure 5.
Effects of antinociceptive, antidepressant, and anticonvulsant drugs on impaired USV calls in rats with inflammatory and neuropathic pain. (A-D) Time course effects of local lidocaine (Lido) blockade of BV or CFA injection sites on impaired emission of 22 kHz and 50 kHz spontaneous calls by inflammatory pain. (D, E) Time course effects of intraperitoneal administration of amitriptyline (AMI) and gabapentin (GBP) on impaired emission of 22 kHz and 50 kHz calls by SNI-induced neuropathic pain. Data expressed as mean ± SEM; n = 8 animals per group; *P < 0.05, **P < 0.01, vs Veh. BV, bee venom; CFA, complete Freund adjuvant; USV, ultrasound vocalization.
Figure 6.
Figure 6.
Reversal effects of local lidocaine (Lido) blockade of peripheral inflammatory pain on impaired emission of spontaneous USVs calls in rats. (A-C) Reversal effects of local Lido blockade of BV and CFA injection sites on impaired emission (powers) of 22 kHz and 50 kHz spontaneous calls caused by peripheral inflammatory pain. (D-G) Reversal effects of Lido blockade of BV and CFA injection sites on impaired emission (number and time) of 22 kHz and 50 kHz calls caused by peripheral inflammatory pain. Data expressed as mean ± SEM; n = 8 animals per group; *P < 0.05, **P < 0.01, ***P < 0.001 vs naive or Veh. BV, bee venom; CFA, complete Freund adjuvant; USV, ultrasound vocalization.
Figure 7.
Figure 7.
Reversal effects of antidepressant and anticonvulsant treatment of neuropathic pain on impaired emission of spontaneous USVs calls in rats. (A, B) Reversal effects of amitriptyline (AMI) and gabapentin (GBP) on impaired emission (powers) of 22 kHz and 50 kHz spontaneous calls caused by SNI-induced neuropathic pain. (C-F) Reversal effects of AMI and GBP on impaired emission (number and time) of 22 kHz and 50 kHz calls caused by SNI-induced neuropathic pain. Data expressed as mean ± SEM; n = 8 animals per group; *P < 0.05, **P < 0.01, ***P < 0.001 vs naive or Veh. SNI, spared nerve injury; USV, ultrasound vocalization.
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