Therapeutic Ultrasound as a Potential Male Dog Contraceptive: Determination of the Most Effective Application Protocol

Canine overpopulation has become a global problem with significant public health impacts. Current population control methods are expensive and/or have questionable efficacy, with continued reliance on euthanasia of large numbers of animals. Male dog contraceptive approaches should be long‐acting or irreversible, highly effective and safe; produce few or no side effects; require limited or no need for significant action to be applied; necessitate no continuing treatment; and be low cost (Verstegen 2010). Other desirable qualities would include no need for invasive surgical procedures or hormonal treatments. Ultrasound (U/S) is a promising canine contraceptive. U/S consists of inaudible high frequency (greater than 20 kHz) mechanical vibrations (pressure waves). The waves are transmitted by propagation through molecular collision and vibration, with a progressive loss of the intensity of the energy during passage through the tissue. When absorbed by tissue, U/S energy waves are converted to heat and it is very effective in deep heating. The temperature of the scrotal testis of mammals is normally several degrees (3–4°C) below that of the body core. An elevation of testicular temperature results in impairment of spermatogenesis (Jung and Schuppe 2007). Thus, it is possible to affect sperm production by raising testicular temperature. Another effect of U/S is mechanical. U/S markedly alters the permeability of the membrane to ions and other substances: U/S increased the sodium concentration and decreased the potassium concentration in the fluid of the seminiferous tubules and decreased the sodium concentration and increased the potassium concentration in the fluid of the rete testis (Fahim 1978). Substances passing from the blood into the seminiferous tubules must pass through the walls of the testicular capillaries, through the tubular wall or into the rete testis and then into the tubular lumen (Setchell 1974). Transport through the capillary wall is passive but is comparatively unrestricted. In contrast, after U/S, there is great variation in the rate at which substances enter the seminiferous tubules and, for some substances, specific transport systems may be involved. These actions have been shown to impact spermatogenesis. Early research on the use of U/S as a suppressant of spermatogenesis found it to be effective even with a small increase in temperature because of its combined thermal and mechanical effects (Fahim et al. 1977). Ultrasound's potential as a male dog contraceptive was first reported in 1975 (Fahim et al. 1975). In a series of publications in the 1970s by Fahim and colleagues, it was shown that a single application of U/S to the testes could result in a dramatic loss of germ cells that was reversible. No notable side effects other than infertility were reported during studies with rats, dogs and monkeys (Fahim et al. 1977). This method was tested on several human subjects, and the authors reported that U/S is a pain‐free procedure that only creates a gentle feeling of warmth (Fahim et al. 1977; Fahim 1978, 1980). The literature over the years highlights different effects on fertility among species. From the studies conducted to date, U/S treatment appears to have a transient contraceptive effect in human and non‐human primates (VandeVoort and Tollner 2012), while in rats (Tsuruta et al. 2012) and dogs (Leoci et al. 2009, 2011), U/S seems to be permanent at the dosages used. Apart from testicular size, frequency and duration also varied among studies. Leoci reported that in dogs treated with three applications of U/S at an intensity setting of 1.5 Watt/cm2, a treatment frequency of 1 MHz on alternating days for 5 min on each testicle was required to obtain suppression of spermatogenesis, but no histological examination was performed. Tsuruta reported that in rats treated with one application of 10 min, treatment frequency of 1 MHz or 3 MHz, and intensity setting of 1–2.2 W/cm2, therapeutic U/S was capable of inducing a nearly complete loss of germ cells. VandeVoort treated four monkeys at 2.5 W/cm2 for 30 min every other day for three treatments total, which resulted in a 91% decrease in sperm count when applied through saline but not for application directly to the testicles. Despite previous promising studies, a standard U/S treatment for contraceptive purposes has not yet been identified, which has limited the usefulness of this approach. Identification of the treatment parameters, including the smallest number of applications needed, the shortest interval among applications, the most effective area of the testis to treat, the speed of the sound head, and the ultrasonic variables (frequency, intensity, duty cycle, etc.), to produce durable contraception has yet to be determined. Thus, the objective of this study was to determine the optimal treatment regime for permanent canine sterilization and identify the nature of testicular tissue damage. Our focus was to find a method that was effective yet easy to perform by investigating various combinations of treatment duration, application number, intensity and frequency parameters.